Document Number: 1

The use of genetically modified organisms (GMOs) as food products
becomes more and more widespread. The European Union has implemented a
set of very strict procedures for the approval to grow, import and/or
utilize GMOs as food or food ingredients. Thus, analytical methods for
detection of the GMOs are necessary in order to verify compliance with
labeling requirements. There are few effective screening methods for
highly processed GM (genetically modified) products. Four genes
(CP4-EPSPS, Cry1A(b), BAR, and, PAT) are common exogenous genes used in
commercialized transgenic soybean, maize, and rice. In the present
study, a multiplex nested polymerase chain reaction (PCR) method was
developed to simultaneously detect the four exogenous genes and one
endogenous gene in two runs. We tested eleven representative highly
processed products samples (soya lecithin, soya protein powder,
chocolate beverage, infant rice cereal, soybean refine oil, soybean
salad oil, maize oil, maize protein powder, maize starch, maize jam)
using the developed method, and amplicons of endogenous gene and
transgenic fragments were obtained from all the processed products
except for soybean refined oil, soybean salad oil and maize oil, and the
sensitivity was 0.005%. These results indicate that multiplex nested PCR
is appropriate for qualitative detection of transgenic soybean, maize
and rice in highly processed products except for refined oil. (C) 2011
Elsevier Ltd. 

Document Number: 2

Substantial equivalence is a key concept in the evaluation of unintended
and potentially harmful metabolic impact consequent to a genetic
modification of food. The application of unsupervised multivariate data
analysis to the metabolic profiles is expected to improve the
effectiveness of such evaluation. The present study uses NMR spectra of
hydroalcoholic extracts, as holistic representations of the metabolic
profiles of grapes, to evaluate the effect of the insertion of one or
three copies of the DefH9-iaaM construct in plants of Silcora and
Thompson Seedless cultivars. The comparison of the metabolic profiles of
transgenic derivatives with respect to their corresponding natural lines
pointed out that the overall metabolic changes occur in the same
direction, independent of the host genotype, although the two cultivars
are modified to different extents. A higher number of copies not only
produces a larger effect but also modifies the whole pattern of
perturbed metabolites.

Document Number: 3

Mycotoxins are secondary metabolites produced by fungi. Mycotoxins in
feed are considered the second most serious issue in the poultry
industry, following increased feed prices. There are different types of
mycotoxins, including aflatoxins, ochratoxins, fumonisin,
trichothecenes, zearalenone, patulin, but the most important are
aflatoxins and ochratoxins. Ochratoxin is known as a nephrotoxin as it
damages kidney tissues predominantly, whereas aflatoxin has more adverse
effects upon the liver. Diagnosis and determination of toxins can be
done in tissues and feed samples by using different techniques such as
high performance liquid chromatography (HPLC), thin layer liquid
chromatography (TLC), and fluorescence polarization immunoassay (FPIA).
Treatment of mycotoxicosis can be done by decontamination or
detoxification of feed by using mycotoxin binders, plant preparations
(nutraceuticals) and vitamins. Mycotoxicosis can be controlled by using
transgenic crops and improving harvesting management, storing feed in
good environmental conditions and fresh feed for the birds. Hazard
analysis critical control point (HACCP) should be referred to for the
control of mycotoxins in the food chain.

Document Number: 4

Crop disease remains a major cause of yield loss and emerging diseases
pose new threats to global food security. Despite the dearth of
commercial development to date, progress in using our rapidly expanding
knowledge of plant-pathogen interactions to invent new ways of
controlling diseases in crops has been good. Many major resistance genes
have now been shown to retain function when transferred between species,
and evidence indicates that resistance genes are more effective when
deployed in a background containing quantitative resistance traits. The
EFR pattern-recognition receptor, present in only the Brassicaceae,
functions to provide bacterial disease control in the Solanaceae.
Knowledge of how transcription activator-like effectors bind DNA is
leading to new methods for triggering disease resistance and broader
applications in genome engineering.

Document Number: 5

Aquaculture is the fastest growing animal producing sector in the world
and is expected to play an important role in global food supply. Along
with this growth, concerns have been raised about the environmental
effects of escapees and pollution, fish welfare, and consumer health as
well as the use of marine resources for producing fish feed. In this
paper we present some of the major challenges salmon farming is facing
today. We discuss issues of relevance to how to ensure sustainability,
by focusing on animal production systems, breeding approaches, sources
for feed ingredients, and genetic engineering strategies. Other crucial
issues such as animal welfare, environmental quality, and ethics are
elaborated with regard to relevance for the sustainability of
aquaculture. Additionally, we comment on socio-economic distributive
implications by intellectual property rights (IPR) strategies on access
to genetic material and traceability. To improve sustainability of
salmon farming we suggest that there is a need for new approaches to
guide research, for identification of ethical issues, and for engaging
stakeholders in resolving these challenges.

Document Number: 6

Since decades, plant male sterility is considered a powerful tool for
biological containment to minimize unwanted self-pollination for hybrid
seed production. Furthermore, prevention of pollen dispersal also
answers to concerns regarding transgene flow via pollen from Genetically
Modified (GM) crops to traditional crop fields or wild relatives. We
induced male sterility by suppressing endogenous general transcription
factor genes, TAFs, using anther-specific promoters combined with
artificial microRNA (amiRNA) technology (Schwab et al., 2006). The
system was made reversible by the ethanol inducible expression of an
amiRNA-insensitive form of the target gene. We provide proof of concept
in eggplant, a cultivated crop belonging to the Solanaceae family that
includes many important food crops. The transgenic eggplants that we
generated are completely male sterile and fertility can be fully
restored by short treatments with ethanol, confirming the efficiency but
also the reliability of the system in view of open field cultivation. By
combining this system with induced parthenocarpy (Rotino et al., 1997),
we provide a novel example of complete transgene containment in
eggplant, which enables biological mitigation measures for the benefit
of coexistence or biosafety purposes for GM crop cultivation.

Document Number: 7

Besides the intended effects that give a genetically modified (GM) plant
the desired trait, unintended differences between GM and non-GM
comparable plants may also occur. Profiling technologies allow their
identification, and a number of examples demonstrating that unintended
effects are limited and diverse have recently been reported. Both from
the food safety aspect and for research purposes, it is important to
discern unintended changes produced by the transgene and its expression
from those that may be attributed to other factors. Here, we show
differential expression of around 0.40% transcriptome between
conventional rice var. Senia and Senia-afp constitutively expressing the
AFP antifungal protein. Analysis of one-fifth of the regulated sequences
showed that around 35% of the unintended effects could be attributed to
the process used to produce GM plants, based on in vitro tissue culture
techniques. A further similar to 15% were event specific, and their
regulation was attributed to host gene disruption and genome
rearrangements at the insertion site, and effects on proximal sequences.
Thus, only around half the transcriptional unintended effects could be
associated to the transgene itself. A significant number of changes in
Senia-afp and Senia are part of the plant response to stress conditions,
and around half the sequences for which up-regulation was attributed to
the transgene were induced in conventional (but not transgenic) plants
after wounding. Unintended effects might, as such, putatively result in
widening the self-resistance characteristics because of the transgene in
GM plants.

Document Number: 8

Strong increase in the production of genetically modified organisms
(GMOs) observed over the years has led to a consolidation of transgenic
seed industries worldwide. The dichotomy between the evaluated risk and
the perceived risk of transgenic use has defined their level of
acceptability among different global societies. GMOs have been widely
applied to agricultural commodities, among them the Roundup Ready (TM)
(RR (TM)) soybean line GTS 40-3-2 has become the most prevalent
transgenic crop in the world. This variety was developed to confer plant
tolerance against glyphosate-based agricultural herbicide Roundup Ready
(TM). Issues related to detection and traceability of GMOs have gained
worldwide interest due to their increasing global diffusion and the
related socioeconomic and health implications. Also, due to the
widespread use of GMOs in food production, labelling regulations have
been established in some countries to protect the right of consumers and
producers. Besides regulatory demand, consumer concern issues have
resulted in the development of several methods of detecting and
quantifying foods derived from genetically engineered crops and their
raw materials. Polymerase chain reaction (PCR) has been proven to be the
method of choice to detect the presence or absence of the introduced
genes of GMOs at DNA level. The present paper aims to verify whether the
PCR technique can detect RR (TM) soybean seeds among conventional ones
to further certification as non-GM soybean seeds and grains. This
analysis could be accomplished through the development of new
methodology called 'intentional contamination' of soybean conventional
seeds or grains with the respective RR (TM) soybeans. The results show
that the PCR method can be applied with high sensitivity in order to
certify conventional soybean seeds and grains.

Document Number: 9

Industrial chemicals and materials are currently derived mainly from
fossil-based raw materials, which are declining in availability,
increasing in price and are a major source of undesirable greenhouse gas
emissions. Plant oils have the potential to provide functionally
equivalent, renewable and environmentally friendly replacements for
these finite fossil-based raw materials, provided that their composition
can be matched to end-use requirements, and that they can be produced on
sufficient scale to meet current and growing industrial demands.
Replacement of 40% of the fossil oil used in the chemical industry with
renewable plant oils, whilst ensuring that growing demand for food oils
is also met, will require a trebling of global plant oil production from
current levels of around 139 MT to over 400 MT annually. Realisation of
this potential will rely on application of plant biotechnology to (i)
tailor plant oils to have high purity (preferably > 90%) of single
desirable fatty acids, (ii) introduce unusual fatty acids that have
specialty end-use functionalities and (iii) increase plant oil
production capacity by increased oil content in current oil crops, and
conversion of other high biomass crops into oil accumulating crops. This
review outlines recent progress and future challenges in each of these
areas.
Practical applications: The research reviewed in this paper aims to
develop metabolic engineering technologies to radically increase the
yield and alter the fatty acid composition of plant oils and enable the
development of new and more productive oil crops that can serve as
renewable sources of industrial feedstocks currently provided by
non-renewable and polluting fossil-based resources. As a result of
recent and anticipated research developments we can expect to see
significant enhancements in quality and productivity of oil crops over
the coming decades. This should generate the technologies needed to
support increasing plant oil production into the future, hopefully of
sufficient magnitude to provide a major supply of renewable plant oils
for the industrial economy without encroaching on the higher priority
demand for food oils. Achievement of this goal will make a significant
contribution to moving to a sustainable carbon-neutral industrial
society with lower emissions of carbon dioxide to the atmosphere and
reduced environmental impact as a result.

Document Number: 10

As a major source of food, cassava (Manihot esculenta Crantz) is an
important root crop in the tropics and subtropics of Africa and Latin
America, and serves as raw material for the production of starches and
bioethanol in tropical Asia. Cassava improvement through genetic
engineering not only overcomes the high heterozygosity and serious trait
separation that occurs in its traditional breeding, but also quickly
achieves improved target traits. Since the first report on genetic
transformation in cassava in 1996, the technology has gradually matured
over almost 15 years of development and has overcome cassava genotype
constraints, changing from mode cultivars to farmer-preferred ones.
Significant progress has been made in terms of an increased resistance
to pests and diseases, biofortification, and improved starch quality,
building on the fundamental knowledge and technologies related to
planting, nutrition, and the processing of this important food crop that
has often been neglected. Therefore, cassava has great potential in food
security and bioenergy development worldwide.

Document Number: 11

Transgenic phytase maize (Zea mays L.) line BVLA430101 was the first
transgenic maize obtained the security certification in 2009 in China.
However, the construct of the phytase gene expression cassette and the
specific detection method have not been reported yet. In this study, the
phytase gene expression cassette was identified, which include maize
legumin promoter, signal peptide, phytase gene, and maize legumin
terminator. The construct-specific qualitative and quantitative PCR
methods of BVLA430101 maize were established based on the transition of
signal peptide and phytase gene using a maize taxon-specific gene zSSIIb
as the endogenous gene. The detection limit for the conventional
qualitative PCR was 200 haploid genome copies of BVLA430101. The
absolute limit of quantification of the real-time PCR was about 20
haploid genome copies. In addition, two known BVLA430101 contents (5 and
1%) of mixed genomic DNA (V/V) were quantified using the developed
real-time PCR detection system, which indicated that the developed
quantitative method can be employed reliably for transgenic phytase
maize BVLA430101 measurement.

Document Number: 12

P>The ability to manipulate plant fatty acid biosynthesis by using new
biotechnological approaches has allowed the production of transgenic
plants with unusual fatty acid profile and increased oil content. This
review focuses on the production of very long chain polyunsaturated
fatty acids (VLCPUFAs) and the increase in oil content in plants using
molecular biology tools. Evidences suggest that regular consumption of
food rich in VLCPUFAs has multiple positive health benefits. Alternative
sources of these nutritional fatty acids are found in cold-water fishes.
However, fish stocks are in severe decline because of decades of
overfishing, and also fish oils can be contaminated by the accumulation
of toxic compounds. Recently, there is also an increase in oilseed use
for the production of biofuels. This tendency is partly associated with
the rapidly rising costs of petroleum, increased concern about the
environmental impact of fossil oil and the attractive need to develop
renewable sources of fuel. In contrast to this scenario, oil derived
from crop plants is normally contaminant free and less environmentally
aggressive. Genetic engineering of the plastid genome (plastome) offers
a number of attractive advantages, including high-level foreign protein
expression, marker-gene excision and transgene containment because of
maternal inheritance of plastid genome in most crops. Here, we describe
the possibility to improve fatty acid biosynthesis in plastids,
production of new fatty acids and increase their content in plants by
genetic engineering of plastid fatty acid biosynthesis via plastid
transformation.

Document Number: 13

Population growth increases the demand for food and thus leads to
expansion of cultivated land and intensification of agricultural
production. There is a definite limit to both of these options for food
security and their multiple negative effects on the environment
undermine the aim for sustainability. Presently the impact of the Green
Revolution on crop production is levelling off at high yields attained
and even the potential of large scale irrigation programmes and
transgenic crops seem to be limited in view of the expected increase in
demand for food. Moreover, climate change threatens to affect
agricultural production across the globe. Skyfarming represents a
promising approach for food production that is largely environment
independent and therefore immune to climate change. Optimal growing
conditions, shielded from weather extremes and pests are aimed at
raising plant production towards the physiological potential. Selecting
rice as a pioneer crop for Skyfarming will not only provide a staple for
a large part of the global population, but also significantly reduce the
greenhouse gas emission caused by paddy cultivation. Multiplication of
the benefits could be achieved by stacking production floors vertically.
In Skyfarming the crop, with its requirements for optimal growth,
development and production, determines the system's design. Accordingly,
the initial development must focus on the growing environment, lighting,
temperature, humidity regulation and plant protection strategies as well
as on the overall energy supply. For each of these areas potentially
suitable technologies are presented and discussed.

Document Number: 14

Pepper (Capsicum L.) is a nutritionally and economically important crop
that is cultivated throughout the world as a vegetable, condiment, and
food additive. Genetic transformation using Agrobacterium tumefaciens
(agrobacterium) is a powerful biotechnology tool that could be used in
pepper to develop community-based functional genomics resources and to
introduce important agronomic traits. However, pepper is considered to
be highly recalcitrant for agrobacterium-mediated transformation, and
current transformation protocols are either inefficient, cumbersome or
highly genotype dependent. The main bottleneck in pepper transformation
is the inability to generate cells that are competent for both
regeneration and transformation. Here, we report that ectopic expression
of the Brassica napus BABY BOOM AP2/ERF transcription factor overcomes
this bottleneck and can be used to efficiently regenerate transgenic
plants from otherwise recalcitrant sweet pepper (C. annuum) varieties.
Transient activation of BABY BOOM in the progeny plants induced prolific
cell regeneration and was used to produce a large number of somatic
embryos that could be converted readily to seedlings. The data highlight
the utility of combining biotechnology and classical plant tissue
culture approaches to develop an efficient transformation and
regeneration system for a highly recalcitrant vegetable crop.

Document Number: 15

Guar seed gum, consisting primarily of a high molecular weight
galactomannan, is the most cost effective natural thickener, having
broad applications in the food, cosmetics, paper, pharmaceutical and
petroleum industries. The properties of the polymer can potentially be
enhanced by genetic modification. Development of suitable
endosperm-specific promoters for use in guar is desirable for metabolic
engineering of the seed gum. A similar to 1.6 kb guar mannan synthase
(MS) promoter region has been isolated. The MS promoter sequence was
fused with the GUS reporter gene and overexpressed in the heterologous
species alfalfa (Medicago sativa). The potential strength and
specificity of the MS promoter was compared with those of the
constitutive 35S promoter and the seed specific beta-phaseolin promoter.
Quantitative GUS assays revealed that the MS promoter directs GUS
expression specifically in endosperm in transgenic alfalfa. Thus, the
guar MS promoter could prove generally useful for directing
endosperm-specific expression of transgenes in legume species.

Document Number: 16

Seed storage proteins are a major component of mature seeds. They are
utilized as protein sources in foods. We designed seed storage proteins
containing bioactive peptides based on their three-dimensional
structures. Furthermore, to create crops with enhanced food qualities,
we developed transgenic crops producing seed storage proteins with
bioactive peptides. This strategy promises to prevent lifestyle-related
diseases by simple daily food consumption. In this review, we discuss a
strategy to develop transgenic crops to improve human health by advanced
utilization of seed storage proteins.

Document Number: 17

The endosperm of cereal grains is an important resource for both food
and feed. It contains three major types of tissue: starchy endosperm,
the aleurone layer, and transfer cells. To improve grain quality and
quantity using molecular methods, control of transgene expression
directed by distinct temporal and spatial promoter activity is
necessary. To identify aleurone layer-specific and/or transfer
cell-specific promoters in rice, microarray analyses were performed,
comparing the aleurone layer containing transfer cells and the other
reproductive and vegetative tissues. After confirmation by RT-PCR
analysis, we identified two putative aleurone layer and/or transfer
cell-specific genes, AL1 and AL2. The promoter regions of these genes
and beta-glucuronidase (GUS) fusion constructs were stably transformed
into rice. The GUS expression patterns indicated that the AL1 promoter
was active exclusively in the dorsal aleurone layer adjacent to the main
vascular bundle. In rice, transfer cells are differentiated in this
region. Therefore, the promoter of the AL1 gene exhibits transfer
cell-containing region-specific activity. The AL1 gene encodes a
putative anthranilate N-hydroxycinnamoyl/benzoyltransferase. The
promoter of this gene will be useful for enhancing uptake of nutrients
from the mother cells and protecting filial seeds from pathogen attack.

Document Number: 18

Numerous crops have been bred or engineered to increase carotenoid
levels in an effort to develop novel strategies that address vitamin A
deficiency in the developing world. The pioneering work in rice (not
covered in this review) has been followed up in many additional crops,
some of which are staples like rice whereas others are luxury products
whose impact on food security is likely to be marginal. This review
surveys the progress that has been made in carotenoid breeding and
metabolic engineering, focusing on beta-carotene enhancement in crops
other than rice. We ask if these efforts have the potential to address
vitamin A deficiency in developing countries by comparing bioavailable
pro-vitamin A levels in wild type and enhanced crops to determine
whether nutritional requirements can be met without the consumption of
unrealistic amounts of food. The potential impact of carotenoid
enhancement should therefore be judged against benchmarks that include
the importance of particular crops in terms of global food security, the
amount of bioavailable beta-carotene, and the amount of food that must
be consumed to achieve the reference daily intake of vitamin A.

Document Number: 19

Reducing pesticide use can provide growers with direct economic benefits
by decreasing the cost of inputs and increasing net returns. Chemical
pesticides will continue to play a role in pest management for the
future. In many situations, the benefits of pesticide use are high
relative to the risks or there are no practical alternatives. The number
and diversity of biological sources will increase, and products that
originate in chemistry laboratories will be designed for particular
target sites. Innovations in pesticide delivery systems in plants
promise to reduce adverse environmental impacts even further. The
correct use of pesticides can deliver significant socio-economic and
environmental benefits in the form of safe, healthy, affordable food;
and enable sustainable farm management by improving the efficiency with
which we use natural resources such as soil, water and overall land use.
Some alternative methods may be more costly than conventional
chemical-intensive agricultural practices, but often these comparisons
fail to account for the high environmental and social costs of pesticide
use. Genetically engineered organisms that reduce pest pressure
constitute a "new generation" of pest management tools. The use of
transgenic crops will probably maintain or even increase the need for
effective resistance management programmes. However, there remains a
need for new chemicals that are compatible with ecologically based pest
management and applicator and worker safety. Evaluation of the
effectiveness of biocontrol agents should involve consideration of
long-term impacts rather than only short-term yield, as is typically
done for conventional practices. The justifications of government
intervention in the management of pest control include the need to
address the externality problems associated with the human and
environmental health effects of pesticides.

Document Number: 20

Systematic reviews and meta-analyses are now routinely used to assess
both the benefits and risks of medical treatments and drugs. The
application of meta-analyses in the medical world is more
straightforward than in the environmental arena because appropriate
control treatments and measures of impact are more easily agreed on in
medicine. Nonetheless, meta-analysis has great potential not only to
inform specific regulatory decisions in the arena of plant
biotechnology, but also to inform general risk management policy. Within
the realm of genetically modified (GM) crops, meta-analyses provide
quantitative syntheses of the benefits, risks, and information gaps of
these emerging technologies. In this context, several published
meta-analyses have examined the risks of insect resistant crops to
nontarget organisms and the risk of outcrossing due to gene flow from GM
crops. The three most critical challenges when applying meta-analysis to
plant biotechnology risk analysis are: (1) identifying the appropriate
comparison-risk compared to what baseline?; (2) using the meta-analysis
to help identify the best levers for risk management; and (3) using the
meta-analysis to estimate the probability of rare, larger magnitude
hazards that are unlikely to occur during short duration studies.
Meta-analysis is an important tool for risk analysis and risk management
because patterns that might never be captured in individual studies can
emerge from the aggregate collection.

Document Number: 21

Genetic modification is the alteration of genetic material that would
not occur naturally. It can be used as a tool to improve the quality of
foods. Through genetic modification foods can be produced in larger
quantities by increasing its resistance to pests and adverse weather
conditions. Their protein and vitamin content may also be increased thus
making them more nutritious. There are also claims of potential risks
such as possible allergic reactions to modified proteins. To date, no
international consensus has been reached for evaluation of the safety of
genetically modified plants for consumption. Over the last decade, the
safety of genetically modified crops in animal feed or foods for human
consumption has been routinely tested in some countries. Protein- and
DNA-based methods have been developed for detection of genetically
modified organisms. Worldwide legislation now faces questions on the use
and labeling requirements of genetically modified crops and their
derivatives. Still, there are concerns about the safety of genetically
modified crops. Also, genetically modified crops could contain toxic
substances or larger amounts of heavy metals and the crops might not be
substantially equivalent in genome, proteome, and metabolome to
unmodified crops. An additional concern is that contrary to
expectations, genetically modified crops may be less nutritious; for
example, they might contain lower amounts of essential phytoestrogens,
needed to protect against heart disease and cancer. This review will
focus on perspectives of the genetically modified food crops.

Document Number: 22

Maize is one of the most important food crops worldwide and is subject
to many diseases. Genetic engineering offers unique way to improve the
current elite line via introduces single or multiple traits. In this
report, we transformed an elite maize line, Jiao 51, with a balsam pear
class I chitinase gene (Mcchit 1) driven by the maize ubiquitin-1 (ubi1)
promoter. Conditions for callus induction from immature embryos were
optimized using a Jiao 51 by culture on induction medium. The highest
Type-I and Type-II callus induction efficiency of 91.33 and 89.10% were
obtained on N6 media supplemented with 2.5 mg L(-1) 2, 4-D. Both types
of calli grew fast. Type-I callus could easily change to Type-II callus.
Agrobacterium-mediated transformation was used to generate eight
transgenic lines that expressed the chitinase transgene. Assessing the
functionality of the Mcchit1 protein in the transgenic plants was
conducted using an excised leaf challenge assay. The transgenic maize
expressing a balsam pear class I chitinase gene exhibited enhanced
tolerance to Exserohilum turcicum in greenhouse conditions. After 5 days
inoculation, the number of lesions was significantly fewer, and the size
of lesions was significantly smaller compared to the controls.

Document Number: 23

The food safety assessment of new agricultural crop varieties developed
through biotechnology includes evaluation of the proteins introduced to
impart desired traits. Safety assessments can include dietary risk
assessments similar to those performed for chemicals intentionally, or
inadvertently added to foods. For chemicals, it is assumed they are not
degraded during processing of the crop into food fractions. For
introduced proteins, the situation can be different. Proteins are highly
dependent on physical forces in their environment to maintain
appropriate three-dimensional structure that supports functional
activity. Food crops such as corn and soy are not consumed raw but are
extensively processed into various food fractions. During processing,
proteins in corn and soy are subjected to harsh environmental conditions
that drastically change the physical forces leading to denaturation and
loss of protein function. These conditions include thermal processing,
changes in pH, reducing agents, mechanical shearing etc. Studies have
shown that processing of introduced proteins such as enzymes that impart
herbicide tolerance or proteins that control insect pests leads to a
complete loss of functional activity. Thus, dietary exposure to
functionally active proteins in processed food products can be
negligible and below levels of any safety concerns. (C) 2010 Elsevier
Ltd. 

Document Number: 24

Background. The density of minerals and vitamins in food staples eaten
widely by the poor may be increased either through conventional plant
breeding or through the use of transgenic techniques, a process known as
biofortification.
Objective. HarvestPlus seeks to develop and distribute varieties of food
staples (rice, wheat, maize, cassava, pearl millet, beans, and sweet
potato) that are high in iron, zinc, and provitamin A through an
interdisciplinary, global alliance of scientific institutions and
implementing agencies in developing and developed countries.
Methods. In broad terms, three things must happen for biofortification
to be successful. First, the breeding must be successful high nutrient
density must be combined with high yields and high profitability.
Second, efficacy must be demonstrated the micronutrient status of human
subjects must be shown to improve when they are consuming the
biofortified varieties as normally eaten. Thus, sufficient nutrients
must be retained in processing and cooking and these nutrients must be
sufficiently bioavailable. Third, the biofortified crops must be adopted
by farmers and consumed by those suffering from micronutrient
malnutrition in significant numbers.
Results. Biofortified crops offer a rural-based intervention that, by
design, initially reaches these more remote populations, which comprise
a majority of the undernourished in many countries, and then penetrates
to urban populations as production surpluses are marketed. In this way,
biofortification complements fortification and supplementation programs,
which work best in centralized urban areas and then reach into rural
areas with good infrastructure.
Conclusions. Initial investments in agricultural research at a central
location can generate high recurrent benefits at low cost as adapted,
biofortified varieties become available in country after country across
time at low recurrent costs.

Document Number: 25

The 14-16 kDa rice allergenic proteins have been reported to be the
products of a multigene family, and react most frequently with IgE in
serum of rice allergic patients. The aim of this study was to assay the
content of endogenous 14-16 kDa rice major allergenic proteins in both
phytase-transgenic (OR) and non-transgenic rice (NR) (wild-type, Oryza
sative L. cv. Tainung 67). We want to investigate whether the foreign
gene will change the allergenicity of rice or not, should be evaluated
in order to clarify its pre-market safety. First, the RA17 gene encoding
16 kDa rice allergenic protein was amplified by reverse
transcription-polymerase chain reaction (RT-PCR), and cloned into the
expression vector pET-29a(+). The constructed expression vector,
pET-29a(+)-RA17, was transformed into Escherichia coli BL21 (DE3). The
host strain was induced to express recombinant RA17, which was purified
by Ni(2+)-NTA resin column and used as the antigen for the production of
a polyclonal antibody via rabbits. We used the antibody as a detective
tool of western blotting and enzyme-linked immunosorbent assay (ELISA)
methods. The western blotting analysis showed that the content of major
allergenic proteins in phytase-transgenic rice was 96.3% of that in
wild-type rice. The content of 14-16 kDa allergens estimated by ELISA
were 1.73 +/- 0.09 (NR) and 1.66 +/- 0.08 mg/g (OR). The results
indicated that the level of 14-16 kDa allergens was not significantly
different (p > 0.05) between wild-type rice and phytase-transgenic rice,
and the transformation of phytase gene into rice plant did not enhance
the content of rice major allergenic proteins in transgenic rice.

Document Number: 26

Rainbow papaya (Carica papaya L) is a genetically engineered (GE)
cultivar with resistance to papaya ringspot virus (PRSV). This cultivar
currently accounts for about 70% of Hawaii's papaya acreage. The
nutritional composition of Rainbow papaya and a non-transgenic control
were analyzed to address GE food safety concerns regarding the potential
for altered nutritional composition and altered expression of inherent
allergens and toxic proteins. Rainbow papaya fruit were analyzed at
three ripening stages and the data compared to that of a non-transgenic
papaya which shares a similar pedigree. No differences were observed
between GE and non-GE papaya for 36 nutrients at any of the tested fruit
ripeness stages. However, vitamin A was higher and calcium levels were
lower in the GE fruit. The GE fruit showed higher levels of protein and
papain at the earliest stage of ripening (color break), but in ripened
fruit these differences were insignificant. Benzyl isothiocyanate (BITC)
levels were very low and similar for both Rainbow and the non-transgenic
control fruit at all ripeness stages. Our data show that the contents of
nutrients, BITC and papain of GE Rainbow papaya are within the range of
those of non-GE papaya and that the Rainbow cultivar is substantially
similar to the non-GE cultivar. Published by Elsevier Inc.

Document Number: 27

Sorghum (Sorghum bicolor (L.) Moench) is an important source for food,
feed, and possesses many agronomic attributes attractive for a biofuels
feedstock. A warm season crop originating from the semi-arid tropics,
sorghum is relatively susceptible to both cold and freezing stress.
Enhancing the ability of sorghum to tolerate cold and freezing offers a
route to expand the acreage for production, and provides a potential
drought avoidance strategy during flowering, an important parameter for
protection of yield. Targeted perturbation of the signal transduction
pathway, that is triggered by exposure to abiotic stress in plants, has
been demonstrated in model systems as an avenue to augment tolerance.
Calcium-dependent protein kinases (CDPKs) are key players in a plant's
response to environmental assaults. To test the impact of modulating
CDPK activity in sorghum as a means to enhanced abiotic stress
tolerance, we introduced a constitutively expressed rice CDPK-7
(OsCDPK-7) gene construct. Sorghum transformants carrying this cassette,
were not improved in cold or salt stress under the conditions tested.
However, a lesion mimic phenotype and upregulation of a number of
pathogen related proteins, along with transcripts linked to
photosynthesis were observed. These results demonstrate that modulating
the Ca signaling cascade in planta via unregulated enhanced CDPK
activity can lead to off-type effects likely due to the broadly
integrated nature of these enzymes in signaling.

Document Number: 28

Toxicity of cassava arises due to the presence of the cyanoglucosides
linamarin and lotaustralin which are hydrolysed by endogenous enzyme
linamarase to acetonecyanohydrin (ACN) and cyanide (CN) which are toxic.
Major research efforts to eliminate/reduce cyanoglucosides have focused
on (i) development of acyanogenic cassava varieties by breeding; (ii)
controlling its metabolism; and (iii) processing to remove cyanogens.
The cyanoglucoside (CNG) content in cassava is genetically controlled
and cultivars may be classified as low (< 50 mu g/g), medium (50-100 mu
g/g) and high CN (> 100 mu g CN eq./g) varieties. Molecular techniques
for reducing tuber CNG have focused on development of transgenic plants
with reduced expression of cyt P 450 in leaves, or increased expression
of hydroxynitrilelyase in tuber. For immediate solution, CNG content can
be reduced using several processing methods. Traditional methods used
for processing include boiling, drying, parboiling and drying, baking,
steaming, frying and preparation of flour. These processes result in CN
losses ranging from 25% to 98%. The cyanogen level in the final product
is influenced both by the tuber CNG and the method of processing. In
order to achieve safe levels of 10 mu g/g in cassava products, new
methods of processing, especially for cassava containing more than 250
mu g CN eq./g, remains a challenging problem. (c) 2010 Published by
Elsevier Ltd.

Document Number: 29

Rice (Oryza sativa L.), one of the most agronomically important crops,
supplies staple food for more than half of the world's population,
especially those living in developing countries. The intensively
increasing world population has put a great burden on rice production.
Drought as one of the major limiting factors for rice productivity has
challenged researchers to improve both the water management system and
rice characteristics. Biotechnology has assisted researchers to identify
genes that are responsive toward drought. This review consolidates the
recent studies that expose a number of drought-responsive genes in rice,
which are potential candidates for development of improved
drought-tolerant transgenic rice cultivars. In addition, examples are
provided of how various drought-responsive genes, such as transcription
factor and protein kinase encoding genes, were explored to engineer rice
plants for enhanced drought tolerance using transgenic approach.
Furthermore, the involvement of various phytohormones in regulation of
drought response as well as the complexity of drought-responsive
networks, which is indicated by the crosstalks with other
stress-responsive networks such as cold and salt stresses, will be
discussed. It is hoped that by understanding how rice responds to
drought, crop performance can be stabilized and protected under water
deficit conditions.

Document Number: 30

Introduction of artificial neural network (ANN) into the field of GMO
detection is the aim of this investigation. The usefulness of ANN to
predict transgenic maize (Bt-176) based on chemical composition of the
extracted crude oil was evaluated. The training set, comprised of a
composition of major and minor lipid components as inputs and outputs.
Crude oil extracted from the genetically modified maize (Bt-176) and
non-transgenic maize was characterized in terms of its fatty acids,
phytosterols and tocopherols distribution as well as of its lipid
classes and unsaponifiables amounts. The results obtained from lipid
distribution analysis showed that the grains of Bt-176 maize were
comparable in their composition to that of the control maize. The
analytical data have been elaborated by supervised pattern recognition
technique ANN in order to classify genetically modified maize (Bt-176)
and conventional maize as well as to authenticate the origin of the
samples.

Document Number: 31

Legumes, including soybeans (Glycine max), are deficient in
sulfur-containing amino acids, which are required for the optimal growth
of monogastric animals. This deficiency can be overcome by expressing
heterologous proteins rich in sulfur containing amino adds in soybean
seeds. A maize 27 kDa gamma-zein, a cysteine-rich protein, has been
successfully expressed in several crops including soybean, barley, and
alfalfa with the intent to biofortify these crops for animal feed.
Previous work has shown that the maize 27 kDa zein can withstand
digestion by pepsin and elicit an immunogenic response in young pigs. By
use of sera from patients who tested positive by ImmunoCAP assay for
elevated IgE to maize proteins, specific IgE binding to the 27 kDa
gamma-zein is demonstrated. Bioinformatic analysis using the full-length
and 80 amino acid sliding window FASTA searches identified significant
sequence homology of the 27 kDa gamma-zein with several known allergens.
Immunoblot analysis using human serum that cross-reacts with maize seed
proteins also revealed specific IgE-binding to the 27 kDa gamma-zein in
soybean seed protein extracts containing the 27 kDa zein. This study
demonstrates for the first time the allergenicity potential of the 27
IcDa gamma-zein and the potential that this. protein has to limit
livestock performance when used in soybeans that serve as a biofortified
feed supplement.

Document Number: 32

We present an extensive literature review exploring the relationships
between food insecurity and rapid biodiversity loss, and the competing
methods proposed to address each of these serious problems. Given a
large and growing human population, the persistence of widespread
malnutrition, and the direct and significant threats the expanding
agricultural system poses to biodiversity, the goals of providing
universal food security and protecting biodiversity seem incompatible.
Examining the literature shows that the current agricultural system
already provides sufficient food on a worldwide basis, but in doing so
methodically undermines the capacity of agroecosystems to preserve
biodiversity. However, the available evidence emphasizes the
interdependence of biodiversity and agriculture, and the important role
each plays in the maintenance of the other. Thus, our review supports
the claim that the solutions to the problems of widespread food
insecurity and biodiversity loss need not be mutually exclusive, and
that it may be possible to address both using appropriate alternative
agricultural practices.

Document Number: 33

BERGEROVA E., GODALOVA Z., SIEKEL P. (2011): Combined effects of
temperature, pressure and low pH on the amplification of DNA of plant
derived foods. Czech J. Food Sc., 29: 337-345.
The effect of food processing on the DNA integrity was studied by means
of PCR amplification of soybean, transgenic MON 810 and non-transgenic
maize, bean, and pea. The degree of DNA degradation was checked by PCR
and visualised by agarose gel electrophoresis. The conditions of
technological treatment such as temperature, pH, pressure, and their
combination may negatively influence the integrity of DNA in processed
foods and hence PCR detection of food components. The DNA over 300 bp
was amplifiable when mild processing parameters up to 100 C were
performed at approximately neutral or low acidic pH. The autoclaving (12
degrees C; 0.1 MPa) significantly reduced the size of amplifiable DNA in
the time dependant manner and that was intensified by acidic pH. The
maximum amplicons length achieved for highly processed matrices was 300
bp. The major impact on the DNA integrity was exerted by the combination
of pressure, temperature, and low pH.

Document Number: 34

Plant carotenoids are C40 isoprenoids with multiple biological roles.
Breeding for carotenoid content in rice, maize and wheat is a relevant
issue, both for their importance in human health and nutrition and for
their influence in food colouration in products such as pasta from durum
wheat. Regarding human health, vitamin A deficiency (VAD) is one of the
major causes of malnutrition in the world. As many as 500 000 children
become blind due to VAD each year with many of them dying from
VAD-related illness within 1 year.
This review presents the main results in the improvement of endosperm
carotenoid levels in rice, maize and wheat considering the methodology
used, either transgenic or non-transgenic; the breeding target, such as
provitamin A or total carotenoid content; the identification of new
carotenogenic genes/alleles related to the available variation for this
trait; and the development of new functional markers for marker-assisted
selection. A comparative overview among these species and key areas for
further improvement are also identified. Carotenoid enhancement in
grasses would benefit from comparative studies among Triticeae species
since they allow the understanding of the diversity basis. Therefore,
the comparative overview given in this work will be relevant not only to
rice, maize and wheat but also to other Triticeae species.

Document Number: 35

Biotransformation of arsenic includes oxidation, reduction, methylation,
and conversion to more complex organic arsenicals. Members of the class
of arsenite (As(III)) S-adenosylmethyltransferase enzymes catalyze
As(III) methylation to a variety of mono-, di-, and trimethylated
species, some of which are less toxic than As(III) itself. However, no
methyltransferase gene has been identified in plants.
Here, an arsM gene from the soil bacterium Rhodopseudomonas palustris
was expressed in Japonica rice (Oryza sativa) cv Nipponbare, and the
transgenic rice produced methylated arsenic species, which were measured
by inductively coupled plasma mass spectrometry (ICP-MS) and
high-performance liquid chromatography-inductively coupled plasma mass
spectrometry (HPLC-ICP-MS).
Both monomethylarsenate (MAs(V)) and dimethylarsenate (DMAs(V)) were
detected in the roots and shoots of transgenic rice. After 12 d exposure
to As(III), the transgenic rice gave off 10-fold greater volatile
arsenicals.
The present study demonstrates that expression of an arsM gene in rice
induces arsenic methylation and volatilization, theoretically providing
a potential stratagem for phytoremediation.

Document Number: 36

Studies suggest a diet rich in long chain omega-3 fatty acids (LCn3) can
promote visual, neural and vascular health. Pork muscle typically has
small amounts of LCn3. A transgenic technology involving the fat-1 gene
has been developed to increase LCn3 in various pig tissues; however,
pork loin muscle has not been examined to date. We have determined that
the LCn3 content in loin muscle (Longissimus) from fat-1 pigs was
1.59-2.24 g/100 g lipid compared to 0.34-0.38 g/100 g lipid in control
samples. Lipid oxidation products were measured in ground and salted
loin muscle from control and fat-1 pigs during storage at -4 degrees C.
There was no clear trend as to whether the fat-1 gene technology
increased or decreased oxidative stability of the muscle during storage
based on lipid peroxide and thiobarbituric acid reactive substances
(TBARS) values. There were a greater number of volatiles detected in
stored loin from fat-1 pigs compared to control. Volatiles common to
control and fat-1 pigs were elevated in stored loin from fat-1 pigs.
This is the first report indicating that the use of fat-1 transgenic
technology significantly increases the amount of desirable LCn3 in pork
loin muscle.

Document Number: 37

While torenia (Torenia fournieri Lind.) is a useful model flower for
molecular biological studies of floral architecture, the maintenance of
plant materials and resulting transgenic plants requires vegetative
propagation due to its heterozygous nature. Reduction of labor and costs
for maintaining thousands of in vitro torenia cultures is therefore a
critical issue. We found that substituting trehalose for sucrose
drastically extended the culture period to 70 days, which is more than
twice as long as for the common, sucrose-based medium, without reduction
in plant viability. Comparative measurement of the plant mass indicated
that the increased survival benefit of the trehalose-based medium might
be on account of improvement in the rhizosphere environment through
reduction of root density in the culture, rather than by reduced plant
growth. No harmful effects arising from the trehalose-based medium were
observed in 1,800 laboratory lines during the bimonthly subculture for
over 12 months, except for a wilting on the first transfer to the
trehalose-based medium. In conjunction with the use of the commercial
food additive, Okome-ni-TREHA (R) rather than reagent-grade trehalose,
we have succeeded in reducing the costs and labor associated with the
culture medium to less than one third of those for the sucrose-based
system.

Document Number: 38

While torenia (Torenia fournieri Lind.) is a useful model flower for
molecular biological studies of floral architecture, the maintenance of
plant materials and resulting transgenic plants requires vegetative
propagation due to its heterozygous nature. Reduction of labor and costs
for maintaining thousands of in vitro torenia cultures is therefore a
critical issue. We found that substituting trehalose for sucrose
drastically extended the culture period to 70 days, which is more than
twice as long as for the common, sucrose-based medium, without reduction
in plant viability. Comparative measurement of the plant mass indicated
that the increased survival benefit of the trehalose-based medium might
be on account of improvement in the rhizosphere environment through
reduction of root density in the culture, rather than by reduced plant
growth. No harmful effects arising from the trehalose-based medium were
observed in 1,800 laboratory lines during the bimonthly subculture for
over 12 months, except for a wilting on the first transfer to the
trehalose-based medium. In conjunction with the use of the commercial
food additive, Okome-ni-TREHA(R) rather than reagent-grade trehalose, we
have succeeded in reducing the costs and labor associated with the
culture medium to less than one third of those for the sucrose-based
system.

Document Number: 39

Rice the major staple food crop which feeds more than half of the
world's population but, lacks pathway to synthesize and accumulate
provitamin A in endosperm therefore rice eaters particularly children,
and pregnant women suffer due to vitamin A deficiency. The pathway for
provitamin A synthesis in rice endosperm has been engineered and
transgenic rice lines have yellow endosperm, called 'Golden Rice'. The
present study aimed at studying the inheritance of transgene(s) in six
transgenic events of 'Golden Rice' and transfer of provitamin A trait
from transgenic lines to a widely grown mega rice variety Swama. The
events E1, R1 and W1 showed normal Mendelian inheritance in F(2),
BC(1)F(1) and BC(1)F(2) generations. The event W1 was studied in
BC(1)F(3) as well and showed normal Mendelian inheritance of 3:1. The
inheritance pattern in L1 event in BC(1)F(1) and BC(1)F(2) showed normal
Mendelian inheritance following expected ratio 1:1 and 3:1 respectively.
The two events G1 and T1 showed distorted segregation in BC(1)F(2) and
BC(2)F(2) respectively in Swama genetic background. In GI event,
transgene inheritance showed segregation distortion in BC(1)F(2) in
favour of transgene negative plants. In T1 event, inheritance followed
expected Mendelian segregation in BC(1)F(1), BC(2)F(1) and BC2F2,
generations. However, when tested against co-dominant inheritance 1:2:1
pattern in BC2F2, segregation distortion was observed with less than the
expected transgene homozygotes. While against 3:1 ratio, it showed the
expected segregation pattern in BC(2)F(2) generation. Segregation
distortion probably due to differential transmission of transgene
positive/negative gametes through either/both parents which needs
further study.

Document Number: 40

Crop varieties with multiple GM events combined by conventional breeding
have become important in global agriculture. The regulatory requirements
in different countries for such products vary considerably, placing an
additional burden on regulatory agencies in countries where the
submission of additional data is required and delaying the introduction
of innovative products to meet agricultural needs. The process of
conventional plant breeding has predictably provided safe food and feed
products both historically and in the modern era of plant breeding.
Thus, previously approved GM events that have been combined by
conventional plant breeding and contain GM traits that are not likely to
interact in a manner affecting safety should be considered to be as safe
as their conventional counterparts. Such combined GM event crop
varieties should require little, if any, additional regulatory data to
meet regulatory requirements. (c) 2010 Elsevier Ltd. All rights
reserved.

Document Number: 41

Pests and diseases impact on crop yield and quality, and also reduce
resource-use efficiency. Improved crop protection strategies to prevent
such damage and loss can increase production and make a substantial
contribution to food security. DNA-based technologies are likely to
greatly increase the speed, sensitivity and accuracy of pest and
pathogen detection and diagnosis. Rapid sequencing of nucleic acids from
infected plants will aid identification of novel disease agents.
Biomarkers of disease or crop damage such as volatile chemicals or
blends may also be used to detect pest outbreaks. Biosensors coupled to
information networks will provide real-time monitoring and surveillance
of crops or stored produce and hence early warning of emerging problems
and new invasive species. Challenges remain in the dissemination of new
technologies and information to resource poor farmers in developing
countries, although the rapid extension of the internet, mobile phones
and other communication networks will provide new opportunities.
Defining the genetic and molecular basis of innate plant immunity has
been a major advance in plant biology with the potential to identify new
targets for intervention via novel chemistry or genetic modification
(GM). Identification of regulatory genes, signal molecules, pathways and
networks controlling induced plant defence should lead to the
development of a new generation of defence modulators, delivered either
as crop protection products, or via biological agents on seeds or in the
root zone. There should also be opportunities to select more responsive
crop genotypes, or to develop transgenic crops tailored to respond to
specific chemical cues or molecular patterns diagnostic for particular
biotic threats. Sequencing of the genomes of the major crop species and
their wild relatives will expand enormously the known gene pool and
diversity of genetic resources available for plant breeders to access.
It should be possible to identify genomic regions and genes conferring
more durable, quantitative resistance to pathogens. The breeding cycle
will be accelerated by high-throughput phenotyping and more efficient
selection of resistance traits using within-gene markers. GM approaches
will facilitate pyramiding (combining) resistance genes with different
specificities and modes of action, thereby reducing the risk of
directional selection for virulence. Analysis of the genomes of plant
pathogens and invertebrate pests is already providing new information on
genes, gene families and processes involved in host colonization and
pathogenicity. Comparative genomics of species with diverse host ranges,
contrasting feeding habits and different pathogenic lifestyles will
identify new targets for inhibiting pest attack and aid the development
of novel antimicrobial drugs and pesticides. Understanding the natural
ecology of pests and pathogens, such as the factors determining host
location, resource exploitation and interactions with other organisms,
will improve our ability to manipulate behaviour, or exploit natural
enemies or other antagonists of pest species. Volatile signals, either
from natural plant sources, or engineered in transgenic crops, will be
more widely used to modify pest behaviour. It may also be possible to
manipulate microbial communities regulating pathogen populations and
activity, and thereby recruit and retain more effective biocontrol
agents.
Insights into the natural diversity and activity of soil and microbial
populations in the zones surrounding roots and seeds will provide new
information on mechanism of suppression regulating pest species. Fully
effective interventions are unlikely, due to the complexity and
diversity of the soil system, but there should be progress towards
integrated control regimes combining more resistant crop genotypes
(either selected or GM) with targeted management of natural suppressive
processes. Harnessing new technologies and knowledge to create more
durable resistant crops and sustainable disease and pest management
systems will require improved understanding of the factors driving pest
and pathogen adaptation and evolution. There must also be an increased
emphasis on translational research and delivery, and developing
strategies appropriate for lower-input production systems, if the second
'green revolution' is to become a reality.

Document Number: 42

Detection of chemical compounds in food sources is based on the
activation of 7 transmembrane gustatory receptors (GRs) in mammals and
in insects such as Drosophila, although the receptors are not conserved
between the classes. Different combinations of Drosophila GRs are
involved in the detection of sugars, but the activated signaling
cascades are largely unknown. Because 7 transmembrane receptors usually
couple to G-proteins, we tried to unravel the intracellular signaling
cascade in taste neurons by screening heterotrimeric G-protein mutant
flies for gustatory deficits. We found the subunit Go alpha to be
involved in feeding behavior and cell excitability by different
transgenic and pharmacological approaches. Go alpha is involved in the
detection of sucrose, glucose, and fructose, but not with trehalose and
maltose. Our studies reveal that Go alpha plays an important role in the
perception of some sweet tastants. Because the perception of other sweet
stimuli was not affected by mutations in Go alpha, we also found strong
indication for the existence of multiple signaling pathways in the
insect gustatory system.

Document Number: 43

Folates are essential cofactors for one-carbon transfer reactions and
are needed in the diets of humans and animals. Because plants are major
sources of dietary folate, plant folate biochemistry has long been of
interest but progressed slowly until the genome era. Since then,
genome-enabled approaches have brought rapid advances: We now know (a)
all the plant folate synthesis genes and some genes of folate turnover
and transport, (b) certain mechanisms governing folate synthesis, and
(c) the subcellular locations of folate synthesis enzymes and of folates
themselves. Some of this knowledge has been applied, simply and
successfully, to engineer folate-enriched food crops (i.e.,
biofortification). Much remains to be discovered about folates, however,
particularly in relation to homeostasis, catabolism, membrane transport,
and vacuolar storage. Understanding these processes, which will require
both biochemical and -omics research, should lead to improved
biofortification strategies based on transgenic or conventional
approaches.

Document Number: 44

Abiotic stress is the primary cause of crop plant yield losses
worldwide. Drought and salinity stress are the major environmental
challenges faced by agriculture. Improving yield production and
stability under stressful environments is needed to fulfil the food
demand of the ever-growing world population. Numerous genes associated
to plant response(s) to drought and salinity stress have been identified
and characterized, in most cases, in the model plant Arabidopsis.
However, while many of these genes are potential candidates for
improving tolerance to abiotic stress, only a small proportion were
transferred into crop plants. Further, transgenic crop plants
overexpressing the genes of interest were, in most cases, tested under
artificial conditions in the laboratory or controlled greenhouse. Thus,
while many reports on drought and salinity tolerance in transgenic
plants have been published, there is urgent need to test these traits
under field conditions. In this chapter, we discuss recent advances in
engineering drought and salinity tolerance in crop plants with emphasis
on yield and the needs to close the gaps between the laboratory and the
field conditions.

Document Number: 45

Some popular processed foods including French fries contain small
amounts of toxic acrylamide. Efforts to lower the accumulation of this
reactive compound by modifying the production process have a negative
effect on sensory characteristics and are not broadly applicable. This
study optimized a method developed more than a decade ago to lower the
accumulation of the acrylamide precursors glucose and fructose in
cold-stored tubers. In contrast to the original application, which
lowered hexose content by one-third through constitutive expression of
an antisense copy of the cold-inducible acid invertase (Inv) gene, the
current approach was based on tuber-specific expression of an
Inv-derived inverted repeat. Stored tubers of transgenic plants
contained as little as 2% of the reducing sugars that accumulated in
controls. This decline in glucose and fructose formation is
counterbalanced by increased sucrose and starch levels. However, it did
not trigger any phenotypic changes and also did not affect the formation
of free asparagine, ascorbic acid, phenylalanine, and chlorogenic acid.
Importantly, French fries from the low-invertase tubers contained up to
8-fold reduced amounts of acrylamide. Given the important role of
processed potato products in the modern Western diet, a replacement of
current varieties with the low-hexose potatoes would reduce the average
daily intake of acrylamide by one-fourth.

Document Number: 46

The present state of knowledge of the phytochemistry of small molecules
isolated from the roots and leaves of cassava Manihot esculenta Crantz
(Euphorbiaceae) is reviewed Cassava roots are an important source of
dietary and industrial carbohydrates mainly eaten as a source of starch
forming the staple food to over 500 million additionally the roots have
value as a raw material for industrial starch production and for animal
feed giving the crop high economic value but It suffers markedly from
post-harvest physiological deterioration (PPD) The hydroxycoumanns
scopoletin and its glucoside scopolin as well as trace quantities of
esculetin and its glucoside esculin are identified from cassava roots
during PPD The biotechnological prospects for cassava are also reviewed
including a critical appraisal of transgenic approaches for crop
improvement together with its use for bioethanol production due to
cassava s efficient ability to fix carbon dioxide into carbohydrate (C)
2010 Elsevier Ltd All rights reserved

Document Number: 47

Two decades have passed since the first maize transformation protocol.
Genetic breeding has been decisive and essential to transform maize a
major food crop worldwide. Biotechnology plays an important role in
plant genetic breeding, particularly for the introduction of novel
traits in order to improve agronomic performance, medical and industrial
applications and food quality. In recent years, the development of
efficient plant regeneration systems in cereal crops and the field of
biotechnology have opened up new opportunities for genetic
transformation of crop plants. Some monocot plants were initially
considered difficult for genetic engineering, primarily due to their
recalcitrance to in vitro regeneration and their resistance to
Agrobacterium. Continuous efforts and studies of different tissues for
regeneration potential, development of various DNA delivery methods, and
optimization of gene expression cassettes have led to the development of
reliable transformation protocols for major cereals, including maize.
Consequently, this research group has focused its attention on maize
transformation mediated by microprojectile bombardment as a device of
DNA delivery into maize cells. This method offers a rapid and simple way
of introgression of candidate genes into cells. However, there are some
points that still need to be studied and improved in order to achieve
appropriate transformation efficiency to optimize the processing
conditions to obtain fertile plants.

Document Number: 48

P>Soybean proteins are the most widely used source vegetable proteins in
the meat industry because of several interesting characteristics. As
soybean is included in the group of ingredients potentially allergenic,
if not declared, it can be considered a hidden allergen, representing a
potential risk to sensitised individuals. The aim of this work was to
optimise and apply DNA-based techniques for soybean detection in meat
products, as alternative to the currently used protein-based methods.
The optimised polymerase chain reaction (PCR) protocol targeting the
soybean lectin gene enabled the detection of the addition of 0.1% and
0.5% of hydrated textured protein, which corresponded to 0.01% and 0.06%
(w/w) of soybean protein in unprocessed and heat-processed pork meats,
respectively. The established PCR technique, when applied to commercial
meat sausages (eighteen samples), confirmed the presence of soybean
declared in nine samples and indicated the presence of soybean in four
samples with no labelled information about soybean. Additionally, the
event-specific PCR detection of Roundup Ready (R) soybean was also
performed, enabling the detection of transgenic DNA in three samples.

Document Number: 49

The struggle over genetically-engineered (GE) maize in Mexico reveals a
deep conflict over the criteria used in the governance of agri-food
systems. Policy debate on the topic of GE maize has become "scientized,"
granting experts a high level of political authority, and narrowing the
regulatory domain to matters that can be adjudicated on the basis of
scientific information or "managed" by environmental experts. While
scientization would seem to narrow opportunities for public
participation, this study finds that Mexican activists acting "in
defense of maize" engage science in multiple ways, using and producing
scientific knowledge as well as treating scientific discussions as a
stage for launching complex social critiques. Drawing from research in
science and technology studies, this article assesses the impacts and
pitfalls of three tactics used by maize activists that respond to the
scientization of biotechnology politics: (1) using scientific
information as a resource; (2) participating in scientific research; and
(3) reframing policy problems as broadly social, rather than as solely
scientific or technical. The obstacles that maize activists have faced
in carrying out each of these efforts indicate that despite diverse and
sophisticated engagements between social movements and the scientific
field, scientization remains a significant institutional barrier to
democratizing agricultural governance.

Document Number: 50

Calcium (Ca) is an essential nutrient for plants and animals, with key
structural and signalling roles, and its deficiency in plants can result
in poor biotic and abiotic stress tolerance, reduced crop quality and
yield. Likewise, low Ca intake in humans has been linked to various
diseases (e.g. rickets, osteoporosis, hypertension and colorectal
cancer) which can threaten quality of life and have major economic
costs. Biofortification of various food crops with Ca has been suggested
as a good method to enhance human intake of Ca and is advocated as an
economically and environmentally advantageous strategy. Efforts to
enhance Ca content of crops via transgenic means have had promising
results. Overall Ca content of transgenic plants has been increased but
in some cases adverse affects on plant function have been observed. This
suggests that a better understanding of how Ca ions (Ca(2+)) are stored
and transported through plants is required to maximise the effectiveness
of future approaches.

Document Number: 51

Seeds of grain legumes are important energy and food sources for humans
and animals. However, the yield and quality of legume seeds are limited
by the amount of sulfur (S) partitioned to the seeds. The amino acid
S-methylmethionine (SMM), a methionine derivative, has been proposed to
be an important long-distance transport form of reduced S, and we
analyzed whether SMM phloem loading and source-sink translocation are
important for the metabolism and growth of pea (Pisum sativum) plants.
Transgenic plants were produced in which the expression of a yeast SMM
transporter, S-Methylmethionine Permease1 (MMP1, YLL061W), was targeted
to the phloem and seeds. Phloem exudate analysis showed that
concentrations of SMM are elevated in MMP1 plants, suggesting increased
phloem loading. Furthermore, expression studies of genes involved in S
transport and metabolism in source organs, as well as xylem sap
analyses, support that S uptake and assimilation are positively affected
in MMP1 roots. Concomitantly, nitrogen (N) assimilation in root and leaf
and xylem amino acid profiles were changed, resulting in increased
phloem loading of amino acids. When investigating the effects of
increased S and N phloem transport on seed metabolism, we found that
protein levels were improved in MMP1 seeds. In addition, changes in SMM
phloem loading affected plant growth and seed number, leading to an
overall increase in seed S, N, and protein content in MMP1 plants.
Together, these results suggest that phloem loading and source-sink
partitioning of SMM are important for plant S and N metabolism and
transport as well as seed set.

Document Number: 52

Environmental stress factors such as drought, elevated temperature,
salinity and rising CO(2) affect plant growth and pose a growing threat
to sustainable agriculture. This has become a hot issue due to concerns
about the effects of climate change on plant resources, biodiversity and
global food security. Plant adaptation to stress involves key changes in
the '-omic' architecture. Here, we present an overview of the
physiological and molecular programs in stress adaptation focusing on
how genes, proteins and metabolites change after individual and multiple
environmental stresses. We address the role which '-omics' research,
coupled to systems biology approaches, can play in future research on
plants seemingly unable to adapt as well as those which can tolerate
climatic change.

Document Number: 53

Plant oil content and composition improvement is a major goal of plant
breeding and biotechnology. The Puroindoline a and b (PINA and PINB)
proteins together control whether wheat seeds are soft or hard textured
and share a similar structure to that of plant non-specific
lipid-transfer proteins. Here we transformed corn (Zea mays L.) with the
wheat (Triticum aestivum L.) puroindoline genes (Pina and Pinb) to
assess their effects upon seed oil content and quality. Pina and Pinb
coding sequences were introduced into corn under the control of a corn
Ubiquitin promoter. Three Pina/Pinb expression positive transgenic
events were evaluated over two growing seasons. The results showed that
Pin expression increased germ size significantly without negatively
impacting seed size. Germ yield increased 33.8% while total seed oil
content was increased by 25.23%. Seed oil content increases were
primarily the result of increased germ size. This work indicates that
higher oil content corn hybrids having increased food or feed value
could be produced via puroindoline expression.

Document Number: 54

Food production and security will be a major issue for supplying an
increasing world population. The problem will almost certainly be
exacerbated by climate change. There is a projected need to double food
production by 2050. In recent times, the trend has been for incremental
modest yield increases for most crops. There is an urgent need to
develop integrated and sustainable approaches that will significantly
increase both production per unit land area and the resource use
efficiency of crops. This review considers some key processes involved
in plant growth and development with some examples of ways in which
molecular technology, plant breeding and genetics may increase the yield
and resource use efficiency of wheat. The successful application of
biotechnology to breeding is essential to provide the major increases in
production required. However, each crop and each specific agricultural
situation presents specific requirements and targets for optimisation.
Some increases in production will come about as new varieties are
developed which are able to produce satisfactory crops on marginal land
presently not considered appropriate for arable crops. Other new
varieties will be developed to increase both yield and resource use
efficiency on the best land.

Document Number: 55

Acid soils limit plant production worldwide because their high
concentrations of soluble aluminium cations (Al(3+)) inhibit root
growth. Major food crops such as wheat (Triticum aestivum L.) have
evolved mechanisms to resist Al(3+) toxicity, thus enabling wider
distribution. The origins of Al(3+) resistance in wheat are perplexing
because all progenitors of this hexaploid species are reportedly
sensitive to Al(3+) stress. The large genotypic variation for Al(3+)
resistance in wheat is largely controlled by expression of an anion
channel, TaALMT1, which releases malate anions from the root apices. A
current hypothesis proposes that the malate anions protect this
sensitive growth zone by binding to Al(3+) in the apoplasm. We
investigated the evolution of this trait in wheat, and demonstrated that
it has multiple independent origins that enhance Al(3+) resistance by
increasing TaALMT1 expression. One origin is likely to be Aegilops
tauschii while other origins occurred more recently from a series of cis
mutations that have generated tandemly repeated elements in the TaALMT1
promoter. We generated transgenic plants to directly compare these
promoter alleles and demonstrate that the tandemly repeated elements act
to enhance gene expression. This study provides an example from higher
eukaryotes in which perfect tandem repeats are linked with
transcriptional regulation and phenotypic change in the context of
evolutionary adaptation to a major abiotic stress.

Document Number: 56

P>Banana Xanthomonas wilt (BXW), caused by the bacterium Xanthomonas
campestris pv. musacearum, is the most devastating disease of banana in
the Great Lakes region of Africa. The pathogen's rapid spread has
threatened the livelihood of millions of Africans who rely on banana
fruit for food security and income. The disease is very destructive,
infecting all banana varieties, including both East African Highland
bananas and exotic types of banana. In the absence of natural host plant
resistance among banana cultivars, the constitutive expression of the
hypersensitivity response-assisting protein (Hrap) gene from sweet
pepper (Capsicum annuum) was evaluated for its ability to confer
resistance to BXW. Transgenic lines expressing the Hrap gene under the
regulation of the constitutive CaMV35S promoter were generated using
embryogenic cell suspensions of two banana cultivars: 'Sukali Ndiizi'
and 'Mpologoma'. These lines were characterized by molecular analysis,
and were challenged with Xanthomonas campestris pv. musacearum to
analyse the efficacy of the Hrap gene against BXW. The majority of
transgenic lines (six of eight) expressing Hrap did not show any
symptoms of infection after artificial inoculation of potted plants in
the screenhouse, whereas control nontransgenic plants showed severe
symptoms resulting in complete wilting. This study demonstrates that the
constitutive expression of the sweet pepper Hrap gene in banana results
in enhanced resistance to BXW. We describe the development of transgenic
banana varieties resistant to BXW, which will boost the arsenal
available to fight this epidemic disease and save livelihoods in the
Great Lakes region of East and Central Africa.

Document Number: 57

Protease inhibitors are a promising complement to Bt toxins for the
development of insect-resistant transgenic crops, but their limited
specificity against proteolytic enzymes and the ubiquity of
protease-dependent processes in living organisms raise questions about
their eventual non-target effects in agroecosystems. After a brief
overview of the main factors driving the impacts of insect-resistant
transgenic crops on non-target organisms, the possible effects of
protease inhibitors are discussed from a multitrophic perspective,
taking into account not only the target herbivore proteases but also the
proteases of other organisms found along the trophic chain, including
the plant itself. Major progress has been achieved in recent years
towards the design of highly potent broad-spectrum inhibitors and the
field deployment of protease inhibitor-expressing transgenic plants
resistant to major herbivore pests. A thorough assessment of the current
literature suggests that, whereas the non-specific inhibitory effects of
recombinant protease inhibitors in plant food webs could often be
negligible and their 'unintended' pleiotropic effects in planta of
potential agronomic value, the innocuity of these proteins might always
remain an issue to be assessed empirically, on a case-by-case basis.

Document Number: 58

Processing of raw plant materials causes occurrence of degraded DNA in
foods. The effect of DNA degradation on amplification and quantification
of transgenic and non-transgenic DNA in raw and experimentally thermally
processed foods was studied. The degree of DNA degradation was checked
by agarose gel electrophoresis and polymerase chain reaction (PCR).
Cetyl trimethyl ammonium bromide method yielded DNA of a better quality,
while Genespin and Wizard were less appropriate. Baking at 220 A degrees
C considerably reduced the size of DNA fragments. In order to measure
the length of amplifiable DNA, primers for soybean and maize genes were
used. Small DNA fragments ranging from 100 to 200 bp were amplified in
all samples. DNA fragments over 1 kbp were amplified only if heating at
220 A degrees C lasted less than 30 min. Baking of flour (220 A degrees
C) reduced the size of extracted DNA fragments so that 1,100 bp amplicon
was no longer amplifiable, while the amplicons of 913 and 1,100 bp were
obtained from the baked bread. When PCR assays targeting maize high
mobility group and zein genes were used under the same conditions,
analogous results were achieved. Quantification of genetically modified
organism content was not influenced by baking.

Document Number: 59

Lovage (Levisticum officinale Koch.), whose leaves and seeds are used to
flavor food, also has medicinal uses. In order to establish a multiple
micropropagation and genetic transformation protocol for lovage, a
reliable and highly efficient method for the regeneration of intact
plants from in vitro culture is essential. In the current study, a rapid
protocol was developed for somatic embryogenesis and plant regeneration
from lovage leaf and stem callus cultures using
2,4-dichlorophenoxyacetic acid (2,4-D). We found that the stem source
was more effective for induction of both embryogenic calluses and
somatic embryos. Nevertheless, in both cases, 2,4-D at 0.1 mg/l was
optimally effective for maximum induction of embryogenic calluses and
somatic embryos per explant. After germination of somatic embryos,
regenerated plantlets were recovered on medium containing half-strength
Murashige and Skoog (MS) salts and vitamins without plant growth
regulator. This system for continuous production of L. officinale
somatic embryos and regenerated plants could be used for
micropropagation of other species as well as for the regeneration of
transgenic plants.

Document Number: 60

Sweet sorghum has substantial potential as a biofuel feedstock, with
advantages in some environments over alternatives such as sugarcane or
maize. Gene technologies are likely to be important to achieve yields
sufficient for food, fuel and fibre production from available global
croplands, but sorghum has proven difficult to transform. Tissue culture
recalcitrance and poor reproducibility of transformation protocols
remain major challenges for grain sorghum, and there has been no
reported success for sweet sorghum. Here we describe a repeatable
transformation system for sweet sorghum, based on (1) optimized tissue
culture conditions for embryogenic callus production with > 90%
regenerability in 12-week-old calli, and (2) an effective selection
regimen for hygromycin resistance conferred by a Ubi-hpt transgene
following particle bombardment. Using this method, we have produced
sixteen independent transgenic lines from multiple batches at an overall
efficiency of 0.09% transformants per excised immature embryo.
Co-expression frequency of a non-selected luciferase reporter was 62.5%.
Transgene integration and expression were confirmed in T(0) and T(1)
plants by Southern analysis and luciferase assays. This success using
the major international sweet sorghum cultivar Ramada provides a
foundation for molecular improvement of sweet sorghum through the use of
transgenes. Factors likely to be important for success with other sweet
sorghum cultivars are identified.

Document Number: 61

Transgenic papaya (Carica papaya L) was produced with the introduction
of replicase (rep) gene for resistance to papaya ringspot virus (PRSV).
In order to investigate the potential unintended compositional changes
in transgenic papaya, profiles of volatile organic compounds (VOCs),
sugar/polyals, organic acids, carotenoids and alkaloids in transgenic
and non-transgenic papaya were obtained respectively by HPLC, GC-MS and
LC-MS, and compared mutually by multivariate statistical methods,
including principal component analysis (PCA) and similarity analysis
method. Results showed that the composition in transgenic papayas
exhibited great similarity to non-transgenic counterparts for measured
components. The contents of important nutrients of beta-carotene and
vitamin C and two natural toxicants, including benzyl isothiocyanate
(BITC) and carpaine, were compared by analysis of variance (ANOVA). The
results also showed that content was similar between transgenic papayas
and non-transgenic counterparts for these components. The variation of
composition in papaya caused by genetic effect was slight during two
harvesting times during our work. It is hoped that this study could
provide some reference value for a safety evaluation of transgenic
papaya from the compositional point of view, and could also propose a
method for discrimination of transgenic food from non-transgenic
counterparts. (C) 2010 Elsevier Inc. 

Document Number: 62

In recent years, strategies and techniques for the production of natural
compounds (plant derived fine chemicals) and/or the breeding of
medicinal and aromatic plants has expanded. Efficient production of high
value natural products with medicinal and cosmetic purpose (e.g.
essential oils, paclitaxel, artemisinin, and vincristine) is the main
target. Metabolic engineering and pathway optimization with the aim to
reduce costs and increase productivity are the main focus of academia
and industry. Until now, only a limited number of plant cell cultures
and isolated enzymes gave sufficiently high production for commercial
purposes. Therefore research strategies have been shifted to metabolic
engineering. Engineering of microorganisms has proven to be a valuable
tool and the concept has been transferred to plant science opening new
promising perspectives. Engineering has been conducted for crop plants,
but the application of this technique to medicinal plants has not yet
been explored well so far. Nowadays the cloning and expression of
multiple genes and genomic integration are of high interest. This allows
the reconstitution of biosynthetic pathways in heterologous organisms,
being either plants or microorganisms. Combining basic science and
applied engineering in this research area has been named combinatorial
biosynthesis and later as synthetic biology. Synthetic biology comprises
a large number of subareas, including enzymology, protein assembly and
interactions, metabolomics, gene regulation, signal transduction and
computational biology and is considered as an important future approach
for biotechnological plant optimization. Synthetic biology has exciting
perspectives for the exploitation of medicinal and aromatic plants, in
order to increase the level of desired natural products, to gain insight
in metabolic pathways even for new similar chemicals, to improve
nutritional and health promoting effects of foods (nutraceuticals), and
to reduce the amount of undesired side products with potential toxic or
allergic activities.

Document Number: 63

P>The branched-chain amino acids (BCAAs) valine, leucine and isoleucine
are essential amino acids that play critical roles in animal growth and
development. Animals cannot synthesize these amino acids and must obtain
them from their diet. Plants are the ultimate source of these essential
nutrients, and they synthesize BCAAs through a conserved pathway that is
inhibited by its end products. This feedback inhibition has prevented
scientists from engineering plants that accumulate high levels of BCAAs
by simply over-expressing the respective biosynthetic genes. To identify
components critical for this feedback regulation, we performed a genetic
screen for Arabidopsis mutants that exhibit enhanced resistance to
BCAAs. Multiple dominant allelic mutations in the VALINE-TOLERANT 1
(VAT1) gene were identified that conferred plant resistance to valine
inhibition. Map-based cloning revealed that VAT1 encodes a regulatory
subunit of acetohydroxy acid synthase (AHAS), the first committed enzyme
in the BCAA biosynthesis pathway. The VAT1 gene is highly expressed in
young, rapidly growing tissues. When reconstituted with the catalytic
subunit in vitro, the vat1 mutant-containing AHAS holoenzyme exhibits
increased resistance to valine. Importantly, transgenic plants
expressing the mutated vat1 gene exhibit valine tolerance and accumulate
higher levels of BCAAs. Our studies not only uncovered regulatory
characteristics of plant AHAS, but also identified a method to enhance
BCAA accumulation in crop plants that will significantly enhance the
nutritional value of food and feed.

Document Number: 64

Background: High amylose starch has attracted particular interest
because of its correlation with the amount of Resistant Starch (RS) in
food. RS plays a role similar to fibre with beneficial effects for human
health, providing protection from several diseases such as colon cancer,
diabetes, obesity, osteoporosis and cardiovascular diseases. Amylose
content can be modified by a targeted manipulation of the starch
biosynthetic pathway. In particular, the inactivation of the enzymes
involved in amylopectin synthesis can lead to the increase of amylose
content. In this work, genes encoding starch branching enzymes of class
II (SBEIIa) were silenced using the RNA interference (RNAi) technique in
two cultivars of durum wheat, using two different methods of
transformation (biolistic and Agrobacterium). Expression of RNAi
transcripts was targeted to the seed endosperm using a tissue-specific
promoter.
Results: Amylose content was markedly increased in the durum wheat
transgenic lines exhibiting SBEIIa gene silencing. Moreover the starch
granules in these lines were deformed, possessing an irregular and
deflated shape and being smaller than those present in the untransformed
controls. Two novel granule bound proteins, identified by SDS-PAGE in
SBEIIa RNAi lines, were investigated by mass spectrometry and shown to
have strong homologies to the waxy proteins. RVA analysis showed new
pasting properties associated with high amylose lines in comparison with
untransformed controls. Finally, pleiotropic effects on other starch
genes were found by semi-quantitative and Real-Time reverse
transcription-polymerase chain reaction (RT-PCR).
Conclusion: We have found that the silencing of SBEIIa genes in durum
wheat causes obvious alterations in granule morphology and starch
composition, leading to high amylose wheat. Results obtained with two
different methods of transformation and in two durum wheat cultivars
were comparable.

Document Number: 65

Indirect modification of animal genomes by interspecific hybridization,
cross-breeding, and selection has produced an enormous spectrum of
phenotypic diversity over more than 10,000 yr of animal domestication.
Using these established technologies, the farming community has
successfully increased the yield and efficiency of production in most
agricultural species while utilizing land resources that are often
unsuitable for other agricultural purposes. Moving forward, animal
well-being and agricultural sustainability are moral and economic
priorities of consumers and producers alike. Therefore, these
considerations will be included in any strategy designed to meet the
challenges produced by global climate change and an expanding world
population. Improvements in the efficiency and precision of genetic
technologies will enable a timely response to meet the multifaceted food
requirements of a rapidly increasing world population.

Document Number: 66

Selection marker genes (SMGs) have been commonly used in genetic
transformation of plants for efficient selection of transformed cells,
tissue, or regenerated shoots. In the majority of cases, the selection
is based on antibiotic or herbicide resistance. The presence of such
genes within the environment or in the food supply might pose an
unpredictable hazard to the ecosystem and to human health; therefore
research has been initiated to develop an efficient marker-free
transgenic system. Various techniques have been developed in recent
years to generate marker free transgenic plants and to eliminate marker
genes from transgenics. These include site-specific recombination,
homologous recombination, transposition, transient co-integration of the
marker gene, and a co-transformation-segregation approach, but success
has been limited to only a few plant species. Transgenic technology
could become more reliable with the improvement of existing marker gene
removal strategies and the development of novel approaches for plant
genome manipulation. This review describes the contemporary strategies
deployed to generate marker-free transgenic plants and marker gene
removal, the merits and shortcomings of different approaches, and
possible directions for future research programmes.

Document Number: 67

New crop varieties containing traits such as enhanced nutritional
profiles, increased yield, and tolerance to drought are being developed.
In some cases, these new traits are dependent on small RNAs or
regulatory proteins such as transcription factors (TF) that modify the
expression of endogenous plant genes. To date, the food and feed safety
of genetically modified (GM) crops has been assessed by the application
of a set of internationally accepted procedures for evaluating the
safety of GM crops. The goal of this paper is to review the main aspects
of the current safety assessment paradigm and to recommend
scientifically sound principles for conducting a safety assessment for
GM crops that are developed by technologies that modify endogenous plant
gene expression. Key considerations for such a safety assessment include
the following:
(1) RNA and TF are generally recognized as safe (GRAS);
(2) Genes encoding RNAi and regulatory proteins such as TFs are an
important component of the plant genome;
(3) Crops engineered using RNAi modifications are not expected to
produce heterologous proteins;
(4) The modulation of TFs may result in quantitative differences in
endogenous plant components, which can be assessed through agronomic
performance and compositional analysis on a case-by-case basis. (C) 2010
ILSI. Published by Elsevier Ltd. 

Document Number: 68

The carotenoids are a major class of organic pigments produced in plants
and microbes. They fulfill many essential physiological and
developmental processes in plants, and also have important roles in
animal health and nutrition. As such they have been the focus of
multidisciplinary research programs aiming to understand how they are
synthesized in microbes and plants, and to clone genes encoding the
corresponding enzymes and express them to modulate carotenoid production
in recombinant microbial and plant systems. Our deeper understanding of
carotenogenic gene regulation, in concert with the development of more
effective multi-gene transfer systems for plants, has facilitated more
ambitious strategies for the modulation of plant carotenoid biosynthesis
not only in laboratory models but more importantly in staple food crops.
Here we review the genetic and molecular tools and resources available
for fundamental and applied carotenoid research, emphasizing recent
achievements in carotenoid engineering and potential future objectives
for carotenoid research in plants. (C) 2010 Elsevier Ireland Ltd. All
rights reserved.

Document Number: 69

Expected increases in world population will continue to make demands on
agricultural productivity and food supply. These challenges will only be
met by increasing the land under cultivation and by improving the yields
obtained on existing farms. Genetic engineering can target key traits to
improve crop yields and to increase production on marginal soils. Soil
acidity is a major abiotic stress that limits plant production
worldwide. The goal of this study was to enhance the acid soil tolerance
of wheat by increasing its resistance to Al(3+) toxicity.
Particle bombardment was used to transform wheat with TaALMT1, the
Al(3+) resistance gene from wheat, using the maize ubiquitin promoter to
drive expression. TaALMT1 expression, malate efflux and Al(3+)
resistance were measured in the T(1) and T(2) lines and compared with
the parental line and an Al(3+)-resistant reference genotype, ET8.
Nine T(2) lines showed increased TaALMT1 expression, malate efflux and
Al(3+) resistance when compared with untransformed controls and null
segregant lines. Some T(2) lines displayed greater Al(3+) resistance
than ET8 in both hydroponic and soil experiments.
The Al(3+) resistance of wheat was increased by enhancing TaALMT1
expression with biotechnology. This is the first report of a major food
crop being stably transformed for greater Al(3+) resistance. Transgenic
strategies provide options for increasing food supply on acid soils.

Document Number: 70

P>The evolution of the seed system provides enormous adaptability to the
gymnosperms and angiosperms, because of the properties of dormancy,
nutrient storage and seedling vigour. Many of the unique properties of
seeds can be exploited in molecular farming applications, particularly
where it is desirable to produce large quantities of a recombinant
protein. Seeds of transgenic plants have been widely used to generate a
raw material for the extraction and isolation of proteins and
polypeptides, which can be processed into valuable biopharmaceuticals.
The factors that control high-level accumulation of recombinant proteins
in seed are reviewed in the following paragraphs. These include
promoters and enhancers, which regulate transcript abundance. However,
it is shown that subcellular trafficking and targeting of the desired
polypeptides or proteins play a crucial role in their accumulation at
economically useful levels. Seeds have proven to be versatile hosts for
recombinant proteins of all types, including peptides or short and long
polypeptides as well as complex, noncontiguous proteins like antibodies
and other immunoglobulins. The extraction and recovery of recombinant
proteins from seeds is greatly assisted by their dormancy properties,
because this allows for long-term stability of stored products including
recombinant proteins and a decoupling of processing from the growth and
harvest cycles. Furthermore, the low water content and relatively low
bioload of seeds help greatly in designing cost-effective manufacturing
processes for the desired active pharmaceutical ingredient. The
development of cGMP processes based on seed-derived materials has only
been attempted by a few groups to date, but we provide a review of the
key issues and criteria based on interactions with Food and Drug
Administration and European Medicines Agency. This article uses 'case
studies' to highlight the utility of seeds as vehicles for
pharmaceutical production including: insulin, human growth hormone,
lysozyme and lactoferrin. These examples serve to illustrate the
preclinical and, in one case, clinical information required to move
these plant-derived molecules through the research phase and into the
regulatory pathway en route to eventual approval.

Document Number: 71

Engineering enhanced transport of zinc to the aerial parts of plants is
a major goal in bio-fortification. In Arabidopsis halleri, high
constitutive expression of the AhHMA4 gene encoding a metal pump of the
P(1B)-ATPase family is necessary for both Zn hyperaccumulation and the
full extent of Zn and Cd hypertolerance that are characteristic of this
species. In this study, an AhHMA4 cDNA was introduced into N. tabacum
var. Xanthi for expression under the control of its endogenous A.
halleri promoter known to confer high and cell-type specific expression
levels in both A. halleri and the non-hyperaccumulator A. thaliana. The
transgene was expressed at similar levels in both roots and shoots upon
long-term exposure to low Zn, control, and increased Zn concentrations.
A down-regulation of AhHMA4 transcript levels was detected with 10 mu M
Zn resupply to tobacco plants cultivated in low Zn concentrations. In
general, a transcriptional regulation of AhHMA4 in tobacco contrasted
with the constitutively high expression previously observed in A.
halleri. Differences in root/shoot partitioning of Zn and Cd between
transgenic lines and the wild type were strongly dependent on metal
concentrations in the hydroponic medium. Under low Zn conditions, an
increased Zn accumulation in the upper leaves in the AhHMA4-expressing
lines was detected. Moreover, transgenic plants exposed to cadmium
accumulated less metal than the wild type. Both modifications of zinc
and cadmium accumulation are noteworthy outcomes from the
biofortification perspective and healthy food production. Expression of
AhHMA4 may be useful in crops grown on soils poor in Zn.

Document Number: 72

Potato (Solanum tuberosum L.) is the world's number one non-cereal food
crop and ranks fourth among most important crops grown worldwide in
terms of acreage, yield and value. In order to maintain greater
protection against environmental stresses, we developed transgenic
potato overexpressing Arabidopsis thaliana glutathione reductase gene
(AtGR1). The transgenic potato maintained up to 6.5 folds higher GR
activity, 5.8 folds glutathione (GSH) contents and up to 2.2 folds
higher glutathione S-transferase activity compared to non transformed
plants (NT). Interestingly, while the transgenic plants exhibited
decreased dehydroascorbate reductase (DHAR) activity, the relative
reduced ascorbate (AsA) contents were higher while the relative
dehydroascorbate (DHA) were lower compared to NT which provide a support
to the hypothesis that an active glutathione-independent pathway for DHA
reduction might exists in vivo. The transgenic plants maintained an
enhanced tolerance to methylviologen, and cadmium. When subjected to
drought stress, the transgenic plants exhibited faster recovery with
less visual injury compared to NT. These results suggest that
manipulation of glutathione levels provides reliable strategy for the
development of industrial transgenic potato plants with enhanced
tolerance to multiple environmental stresses.

Document Number: 73

We developed an event-specific DNA microarray system to identify 19
genetically modified organisms (GMOs), including two GM soybeans
(GTS-40-3-2 and A2704-12), thirteen GM maizes (Bt176, Bt11, MON810,
MON863, NK603, GA21, T25, TC1507, Bt10, DAS59122-7, TC6275, MIR604, and
LY038), three GM canolas (GT73, MS8xRF3, and T45), and one GM cotton
(LLcotton25). The microarray included 27 oligonucleotide probes
optimized to identify endogenous reference targets, event-specific
targets, screening targets (35S promoter and nos terminator), and an
internal target (18S rRNA gene). Thirty-seven maize-containing food
products purchased from South Korean and US markets were tested for the
presence of GM maize using this microarray system. Thirteen GM maize
events were simultaneously detected using multiplex PCR coupled with
microarray on a single chip, at a limit of detection of approximately
0.5%. Using the system described here, we detected GM maize in 11 of the
37 food samples tested. These results suggest that an event-specific DNA
microarray system can reliably detect GMOs in processed foods.

Document Number: 74

P>The aim of this study was to evaluate the use of four nontargeted
analytical methodologies in the detection of unintended effects that
could be derived during genetic manipulation of crops. Three profiling
technologies were used to compare the transcriptome, proteome and
metabolome of two transgenic maize lines with the respective control
line. By comparing the profiles of the two transgenic lines grown in the
same location over three growing seasons, we could determine the extent
of environmental variation, while the comparison with the control maize
line allowed the investigation of effects caused by a difference in
genotype. The effect of growing conditions as an additional
environmental effect was also evaluated by comparing the Bt-maize line
with the control line from plants grown in three different locations in
one growing season. The environment was shown to play an important
effect in the protein, gene expression and metabolite levels of the
maize samples tested where 5 proteins, 65 genes and 15 metabolites were
found to be differentially expressed. A distinct separation between the
three growing seasons was also found for all the samples grown in one
location. Together, these environmental factors caused more variation in
the different transcript/protein/metabolite profiles than the different
genotypes.

Document Number: 75

Genetically modified (GM) foods must be tested for safety, including by
allergenicity tests to ensure that they do not contain new allergens or
higher concentrations of known allergens than the same non-GM foods.
In this study experimentally developed EGFP-transgenic chickens were
used and evaluated the allergenicity of meat from the chicken based on a
serological and two-dimensional difference gel electrophoresis (2D-DIGE)
analysis. For the serological analysis, a Western blotting with
allergen-specific antibodies and a proteomic analysis of chicken meat
allergens with patients' sera, a so-called allergenome analysis, were
used. The allergenome analysis allowed us to identify five IgE-binding
proteins in chicken meat, including a known allergen, chicken serum
albumin, and no qualitative difference in their expressions between the
GM and non-GM chicken meat was found. Results of the 2D-DIGE analysis
showed that none of the IgE-binding proteins in chicken meat were
significantly changed in expression levels between non-GM and GM
chicken, and only 3 of the 1500 soluble protein spots including green
fluorescence protein were markedly different as a result of gene
transfer.
These above results showed that the combination of serological and
2D-DIGE analysis is a valid method of evaluating quality and quantity of
allergens in GM foods. (C) 2010 Elsevier Ltd. 

Document Number: 76

Drought is the most important environmental stress affecting agriculture
worldwide. Exploiting yield potential and maintaining yield stability of
crops in water-limited environments are urgent tasks that must be
undertaken in order to guarantee food supply for the increasing world
population. Tremendous efforts have been devoted to identifying key
regulators in plant drought response through genetic, molecular, and
biochemical studies using, in most cases, the model species Arabidopsis
thaliana. However, only a small portion of these regulators have been
explored as potential candidate genes for their application in the
improvement of drought tolerance in crops. Based on biological
functions, these genes can be classified into the following three
categories: (1) stress-responsive transcriptional regulation (e.g.
DREB1, AREB, NF-YB); (2) post-transcriptional RNA or protein
modifications such as phosphorylation/dephosphorylation (e.g. SnRK2,
ABI1) and farnesylation (e.g. ERA 1); and (3) osomoprotectant metabolism
or molecular chaperones (e.g. CspB). While continuing down the path to
discovery of new target genes, serious efforts are also focused on
fine-tuning the expression of the known candidate genes for stress
tolerance in specific temporal and spatial patterns to avoid negative
effects in plant growth and development. These efforts are starting to
bear fruit by showing yield improvements in several crops under a
variety of water-deprivation conditions. As most such evaluations have
been performed under controlled growth environments, a gap still remains
between early success in the laboratory and the application of these
techniques to the elite cultivars of staple crops in the field.
Nevertheless, significant progress has been made in the identification
of signaling pathways and master regulators for drought tolerance. The
knowledge acquired will facilitate the genetic engineering of single or
multiple targets and quantitative trait loci in key crops to create
commercial-grade cultivars with high-yielding potential under both
optimal and suboptimal conditions.

Document Number: 77

Analysis of food products consisting of, or produced from, genetically
modified organisms (GMOs) is required to verify compliance with labeling
legislation and to detect any possible unauthorized transgenic crops.
With these goals, 240 samples of soy-derived foodstuffs and 25 samples
of maize-derived foodstuffs were analyzed from 2004 to 2007. All samples
positive for Roundup Ready (R) soybean were quantitatively examined
using the TaqMan (R) GMO 35S Soy kit. In food containing soy, 68 (28.3%)
were shown to contain GM soy, whereas in food containing maize, neither
Bt 176 nor MON 810 maize were found. Quantitative analysis revealed GMO
contents ranging from 0.05 to 1% in 43(63.2%) samples, and more than 1%
in 25 (36.8%) samples. The absolute and relative limits of detection
(LODs) were approximately 10 copies and 0.0125%, respectively, and the
absolute and relative limits of quantification (LOQs) were approximately
40 copies and 0.05%, respectively, suggesting sufficient sensitivity to
quantify genetically modified (GM) materials below and above the legal
threshold of 1%. The presence of GM material in these samples was not
indicated on their labels, indicating that none of these food products
had been appropriately labeled. These results clearly demonstrate the
need for a monitoring program of food products by the Brazilian
regulatory authorities. (C) 2010 Elsevier Inc. 

Document Number: 78

The application of genetic engineering to food animals is often viewed
as a means to further increase animal productivity without regard for
the welfare of the resulting animals. We offer the perspective that, on
the contrary, genetic engineering can, and is, being used to improve
animal welfare in modern production systems. Several examples are cited
from the current work in the field of animal genetic engineering that
should be included in the debate over whether genetically engineered
animals should be used in production agriculture. The current debate has
slowed the advancement of this technology, which could play a key role
in improving animal welfare and sustainability, without considering the
potential benefits.

Document Number: 79

Despite the potential benefits of genetically modified foods (GMF), the
technology has its limitations and is surrounded by controversy. This
review focuses on the recent advances and applications of GM in food
production, the present advantages of GMF as well as its limitations and
challenges in safety assessment evolving from the recent developments.
By summarizing the published information from scattered references in
the literature, this paper attempts to contribute to the literature an
overall understanding as well as the current status evolution of this
field.

Document Number: 80

Oxalate is widely distributed in the plant kingdom. While excess oxalate
in food crops is detrimental to animal and human health, it may play
various functional roles in plants, particularly for coping with
environmental stresses. Understanding its biosynthetic mechanism in
plants, therefore, becomes increasingly important both theoretically and
practically. However, it is still a matter of debate as to what
precursor and pathway are ultimately used for oxalate biosynthesis in
plants. In this study, both physiological and molecular approaches were
applied to address these questions. First, it was observed that when
glycolate or glyoxylate was fed into detached leaves, both organic acids
were equally effective in stimulating oxalate accumulation. In addition,
the stimulation could be completely inhibited by cysteine, a glyoxylate
scavenger that forms cysteine-glyoxylate adducts. To verify the role of
glyoxylate further, various transgenic plants were generated, in which
several genes involved in glyoxylate metabolism [i.e. SGAT
(serine-glyoxylate aminotransferase), GGAT (glutamate-glyoxylate
aminotransferase), HPR (hydroxypyruvate reductase), ICL (isocitrate
lyase)], were transcriptionally regulated through RNAi or
over-expression. Analyses on these transgenic plants consistently
revealed that glyoxylate acted as an efficient precursor for oxalate
biosynthesis in rice. Unexpectedly, it was found that oxalate
accumulation was not correlated with photorespiration, even though this
pathway is known to be a major source of glyoxylate. Further, when GLDH
(L-galactono-1,4-lactone dehydrogenase), a key enzyme gene for ascorbate
biosynthesis, was down-regulated, the oxalate abundance remained
constant, despite ascorbate having been largely reduced as expected in
these transgenic plants. Taken together, our results strongly suggest
that glyoxylate rather than ascorbate is an efficient precursor for
oxalate biosynthesis, and that oxalate accumulation and regulation do
not necessarily depend on photorespiration, possibly due to the
occurrence of the anaplerotic reaction that may compensate for
glyoxylate formation in rice.

Document Number: 81

It is now accepted that omega-3 polyunsaturated fatty acids, especially
eicosapentaenoic acid (EPA; 20:5 Delta 5,8,11,14,17) and docosahexaenoic
acid (DHA, 22:6 Delta 4,7,10,13,16,19) play important roles in a number
of aspects of human health, with marine fish rich in these beneficial
fatty acids our primary dietary source. However, over-fishing and
concerns about pollution of the marine environment indicate a need to
develop alternative, sustainable sources of very long chain
polyunsaturated fatty acids (VLC-PUFAs) such as EPA and DHA. A number of
different strategies have been considered, with one of the most
promising being transgenic plants "reverse-engineered" to produce these
so-called fish oils. Considerable progress has been made towards this
goal and in this review we will outline the recent achievements in
demonstrating the production of omega-3 VLC-PUFAs in transgenic plants.
We will also consider how these enriched oils will allow the development
of nutritionally-enhanced food products, suitable either for direct
human ingestion or for use as an animal feedstuff. In particular, the
requirements of aquaculture for omega-3 VLC-PUFAs will act as a strong
driver for the development of such products. In addition,
biotechnological research on the synthesis of VLC-PUFAs has provided new
insights into the complexities of acyl-channel ling and triacylglycerol
biosynthesis in higher plants. (C) 2009 Elsevier Ltd. All rights
reserved.

Document Number: 82

Peanut (Arachis hypogaea) allergy is one of the most serious food
allergies. Peanut seed protein, Ara h 3, is considered to be one of the
most important peanut allergens. Little is known about the temporal and
spatial regulation mechanism of Ara h 3 during seed development. In this
study, chromatin structure of the Ara h 3 promoter was analyzed to
examine its transcriptional regulation. Analysis of transgenic plants of
Arabidopsis thaliana expressing Arah3: GUS showed that the Ara h 3
promoter could efficiently direct the seed-specific expression of the
GUS reporter gene. Chromatin immunoprecipitation revealed that
nucleosomes were depleted at the core promoter of the Ara h 3 upon full
activation in the late stage of embryo maturation, which was accompanied
by a dramatic decrease of histone acetylation. However, in the early
stage of embryo maturation, histone H3 hyperacetylation at the core
promoter of Ara h 3 was detected. A decrease of histone H3-K9
dimethylation levels at core promoter of Ara h 3 was also observed with
concomitant repression of Ara h 3 in the vegetative tissues. Our results
suggest that the histone modification status of Ara h 3 undergoes
targeted changes including the increase of histone H3 acetylation and
decrease of histone H3-K9 dimethylation in early maturation embryos. In
addition, the loss of histone H3 from the proximal promoter of Ara h 3
is associated with its high expression during late embryo maturation.

Document Number: 83

The cultivation area of genetically modified (CM) crops is increasing
all over the world. Though no land in the Republic of Korea is currently
used for the cultivation of GM crops, GM crop imports for food and
foraging purposes are continuously increasing. This may promote the
unintentional escape of GM crops. This study was conducted to
investigate whether imported GM maize is released into our environment
during the transportation of grain in the Republic of Korea. Based on
PCR analysis, most of the maize grains in the forage products were GM,
and about 50% of the grains were germinated. Monitoring was conducted in
two major grain receiving ports, 15 feed manufacturing plants. and 14
livestock barns in five provinces of the Republic of Korea from July to
September 2007. We found many spilled maize grains around open storage
areas of ports and along truck transportation routes near feed
manufacturing plants. Established maize plants were not found at or
around Incheon port. However, we found 18 established maize plants at
the Gunsan port, 15 of which were GM. We also found eight GM maize
plants around four feed manufacturing plants and in two livestock barns.
Based on the event-specific PCR analysis, three maize events (NK603,
Mon810, and TC1507) were identified. Though several GM maize plants were
found around the port and feed manufacturing plants, most of these
facilities were located inside the industrial park and were far from
cultivated fields, likely rendering the impact of these GM maize on the
natural environments negligible. However, most of the livestock barns
were close to cultivated areas. Moreover, maize plants were cultivated
for food or feed near some livestock barns. This practice may facilitate
gene flow from GM maize to non-GM maize plants. Therefore, continuous
monitoring is necessary to detect the occurrence of CM maize, and
appropriate action should be taken to prevent genetic admixture in our
environment. (C) 2009 Published by Elsevier Ltd.

Document Number: 84

The in vitro pepsin digestion assay is the international standard for
assessing the safety or risk of novel proteins newly produced in
transgenic crops. However, this protocol, based on the degradation of
protein purified from Escherichia coli, has recently been criticized for
problems such as its objective detection limit. Here, we estimated the
digestion stability of the phosphinothricin acetyltransferase (PAT)
protein in soluble proteins as well as from leaf tissue powder in
simulated gastric fluid (SGF). Our line of genetically modified
zoysiagrass carried a single copy of the bar gene, which entered a
chromosomal region not encoding protein. We designated it as Jeju Green
21 (JG21). From total soluble proteins extracted from JG21 leaves,
digestibility of the PAT protein in SGF was examined by enzymatic
assays, sodium dodecyl sulfate-polyacrylamide gel electrophoresis,
Western gel blots, and an immunodetection strip kit. The degradability
of pure PAT protein obtained from E. coli was clearly apparent within at
least 30 s. However, PAT degradation in leaf tissue powder was
significantly delayed, indicating that some matrices in that powder
might have influenced its digestion stability by SGF. Nevertheless,
degradation of the powder (real-life) sample was complete within at
least 5 min, suggesting that this protein produced in JG21 zoysiagrass
can be digested harmlessly in the stomachs of humans or livestock.

Document Number: 85

Fusarium head blight (FHB) is a disease that causes major economic
losses in wheat and barley production worldwide. Contamination of food
with the trichothecene mycotoxin deoxynivalenol (DON) produced by
Fusarium is a major health concern for humans and animals because
trichothecenes are potent cytotoxins of eukaryotic cells. Trichothecene
mycotoxins inhibit translation by targeting ribosomal protein L3 at the
peptidyltransferase center. We previously showed that expression of an
N-terminal fragment of yeast L3 (L3 Delta) in transgenic tobacco plants
reduced the toxicity of DON. Here, we produced transgenic wheat plants
that express the same yeast 13 (L3 Delta) fragment and evaluated their
susceptibility to Fusarium graminearum infection and their ability to
accumulate DON. Following F. graminearum infection in greenhouse tests,
two transgenic wheat lines expressing the highest levels of L3 Delta
showed reductions in disease severity and kernel DON levels, compared to
non-transformed plants. In a field test, a transgenic wheat line with
the highest L3 Delta expression controlled by the maize Ubi1 promoter
had significant reductions in visually scabby kernels and kernel DON
levels. These results demonstrate that expression of a modified form of
the ribosomal protein that is the target of DON can improve FHB
resistance in wheat. (C) 2010 Elsevier Ireland Ltd. 

Document Number: 86

BACKGROUND: The increasing global market of genetically modified (GM)
crops amplifies the potential for unintentional contamination of food
and feed with GM plants. Methods proposed for disposal of crop residues
should be assessed to prevent unintended distribution of GM materials.
Composting of organic material is inexpensive and location-independent.
The objective of this study was to determine the effectiveness of
composting for disposal of GM plants in terms of reducing seed viability
and promoting the degradation of endogenous as well as transgenic DNA.
RESULTS: Duplicate samples of corn kernels, alfalfa leaves, and GM
canola seeds, meal and pellets were sealed in porous nylon bags and
implanted in duplicate 85 000 kg (initial weight) feedlot manure compost
piles. Samples were collected at intervals over 230 days of composing.
Canola seeds and corn kernels were not viable after 14 days of
composting with temperatures in the piles exceeding 50 degrees C. In all
samples, PCR analyses revealed that plant endogenous and transgenic
fragments were substantially degraded after 230 days of composting.
Southern blotting of genomic DNA isolated from canola seeds identified
differences in the persistence of endogenous, transgenic, and bacterial
DNA.
CONCLUSION: Composting GM and non-GM plant materials with manure
rendered seeds non-viable, and resulted in substantial, although not
complete, degradation of endogenous and transgenic plant DNA. This study
demonstrates that composting could be effective for disposing of GM
crops in the event of their inadvertent entry into the food or feed
chain. Copyright (C) 2010 Crown in the right of Canada. Published by
John Wiley & Sons, Ltd

Document Number: 87

Rice, the major food crop of world is severely affected by homopteran
sucking pests. We introduced coding sequence of Allium sativum leaf
agglutinin, ASAL, in rice cultivar IR64 to develop sustainable
resistance against sap-sucking planthoppers as well as eliminated the
selectable antibiotic-resistant marker gene hygromycin
phosphotransferase (hpt) exploiting cre/lox site-specific recombination
system. An expression vector was constructed containing the coding
sequence of ASAL, a potent controlling agent against green leafhoppers
(GLH, Nephotettix virescens) and brown planthopper (BPH, Nilaparvata
lugens). The selectable marker (hpt) gene cassette was cloned within two
lox sites of the same vector. Alongside, another vector was developed
with chimeric cre recombinase gene cassette. Reciprocal crosses were
performed between three single-copy T(0) plants with ASAL- lox-hpt-lox
T-DNA and three single-copy T(0) plants with cre-bar T-DNA. Marker gene
excisions were detected in T(1) hybrids through hygromycin sensitivity
assay. Molecular analysis of T(1) plants exhibited 27.4% recombination
efficiency. T(2) progenies of L03C04(1) hybrid parent showed 25% cre
negative ASAL-expressing plants. Northern blot, western blot and ELISA
showed significant level of ASAL expression in five marker-free T(2)
progeny plants. In planta bioassay of GLH and BPH performed on these
T(2) progenies exhibited radical reduction in survivability and
fecundity compared with the untransformed control plants.

Document Number: 88

According to the EU and Swiss legislation for food, only approved traits
of transgene plants are allowed to be imported and sold to the consumer.
In order to control imports of rice and rice products from retailers,
dealers and importing companies, efficient and reliable methods for the
detection and quantification are a prerequisite. Therefore, a novel
pentaplex real-time polymerase chain reaction system was developed and
validated for the quantitative determination of three genetically
modified rice lines at once. This system simultaneously determines DNA
contents of the phospholipase-gen, a rice species specific gene, the
35S:BAR-construct, as the promotor of different transgene rice lines and
the specific systems for LL62, LL601 and Bt-63-rice (Shanyou63). The
test exhibits a good specificity and sensitivity for the transgenes in
the range of 0.01-1%. It proved its efficiency and reliability in daily
routine. Due to the lack of appropriate reference material for the
Bt-63-rice, a reference oligonucleotide was artificially constructed.
This oligonucleotide proved its applicability in diagnostic analysis.

Document Number: 89

The insect-resistant transgenic rice event TT51-1 (synonym BT63) has
been found illicitly planted and distributed for years although it has
never been approved for commercial cultivation in any country up to now.
The purpose of this study was to establish a detection method that is
specific for this transformation event. The event-specific PCR method
produces an amplicon of 120 basepairs (bp) based on the revealed 3'
junction sequence with a limit of detection (LOD) and a limit of
quantification (LOQ) being approximately 5 and 10 initial template
copies, respectively. Two mixed rice samples with known TT51-1 contents
were used to verify the developed real-time PCR system, from which the
expected results were observed. (C) 2009 Elsevier Ltd. All rights
reserved.

Document Number: 90

The release of the Final Guidance from the US Food and Drug
Administration on the commercialization of genetically engineered
animals has sparked renewed discussion over the ethical, consumer, and
regulatory implications of transgenesis in animal agriculture. Animal
welfare critiques have focused on unexpected phenotypic effects in
animals used in transgenic research, rather than on the health and
welfare implications of the intended productivity enhancement. Unless
breeding goals are redefined to reflect social concerns, the occurrence
and magnitude of undesirable side effects may increase and consumer
confidence in the nascent technology may be undermined.

Document Number: 91

Recombinant protease inhibitors represent useful tools for the
development of insect-resistant transgenic crops, but questions have
been raised in recent years about the impact of these proteins on
endogenous proteases and chemical composition of derived food products
In this study, we performed a detailed compositional analysis of tubers
from potato lines expressing the broad-spectrum inhibitor of Ser and Asp
proteases, tomato cathepsin D inhibitor (SICIDI), to detect possible
unintended effects on tuber composition A compositional analysis of key
nutrients and toxic chemicals was carried out with tubers of
SICDI-expressing and control (comparator) lines, followed by a
two-dimensional gel electrophoresis (2-DE) proteomic profiling of total
and allergenic proteins to detect eventual effects at the proteome level
No significant differences were observed among control and
SICDI-expressing lines for most chemicals assayed, in line with the very
low abundance of SICDI in tubers Likewise, proteins detected after 2-DE
showed no quantitative variation among the lines, except for a few
proteins in some control and test lines, independent of sicidi transgene
expression Components of the patatin storage protein complex and Kunitz
protease inhibitors immunodetected after 2-DE showed unaltered
deposition patterns in SICIDI-expressing lines, clearly suggesting a
null impact of slcdi on the intrinsic allergenic potential of potato
tubers These data suggest, overall, a null impact of slcdi expression on
tuber composition and substantial equivalence between comparator and
SICIDI-expressing tubers despite reported effects on leaf protein
catabolism They also illustrate the usefulness of proteomics as a tool
to assess the authenticity of foods derived from novel-generation
transgenic plants.

Document Number: 92

Genetically modified (GM) plants aimed at producing food/feed are part
of regular agriculture in many areas of the World. Commodity plants have
also found application as bioreactors, designated nonfood/non-feed CIA
(NFGM) plants. thereby making raw material for further refinement to
industrial. diagnostic or pharmaceutical preparations Many among them
may pose health challenge to consurners or livestock animals. if
occurring in food/feed. NFGM plants are typically released into the
environment, but are grown under special oversight and any among several
containment practices, none of which provide full protection against
accidental dispersal. Adventitious admixture with food or feed can Occur
either through distributional mismanagement or as a consequence of gene
flow to plant relatives. To facilitate NFGM surveillance we propose a
new mandatory tagging of essentially all such plants. prior to
cultivation or marketing in the European Union. The suggested tag -
Plant-Made Industrial or Pharmaceutical Products Tag (PMIP-T) - is
envisaged to occur as a transgenic silent DNA identifier in host plants
and designed to enable technically simple identification and
characterisation of any NFGM. Implementation of PMIP-T would permit
inexpensive, reliable and high-throughput screening for NFGM
specifically The paper outlines key NFGM prospects and challenges as
well as the PMIP-T concept. (C) 2009 Elsevier Ltd All rights reserved

Document Number: 93

Anthranilate synthase (AS) is a key regulatory enzyme in tryptophan
(Trp) biosynthesis and is subject to feedback inhibition by Trp. The
gene encoding a mutated feedback-resistant alpha subunit of rice AS
(OASA1D) under the control of either a soybean glycinin gene promoter or
the 35S promoter of cauliflower mosaic virus for seed-specific or
constitutive expression, respectively, was introduced into soybean
[Glycine max (L.) Merrill] by particle bombardment. A total of seven
different transgenic lines that showed markedly increased accumulation
of free Trp in their seeds were developed. The overproduction of free
Trp was stably inherited in subsequent generations without any apparent
detrimental effect on plant growth or reproduction. The total Trp
content of transgenic seeds was also about twice that of nontransgenic
seeds, whereas the amount of protein-bound Trp was not substantially
affected by OASA1D expression. In spite of the marked increase in free
Trp content, metabolic profiling by high-performance liquid
chromatography coupled with mass spectrometry revealed little change in
the amounts of other aromatic compounds in the transgenic seeds. We
developed a rapid and feasible system based on farmed rainbow trout to
evaluate the nutritional quality of a limited quantity of transgenic
soybean seeds. Supplementation of fish food with OASA1D transgenic
soybean seeds or with nontransgenic seeds plus crystalline Trp increased
the growth rate of the farmed fish. These results indicate
transformation with OASA1D is a reliable approach to improve the
nutritional quality of soybean (or of other grain legumes) for human and
animal food.

Document Number: 94

Natural resources and environmental quality are in constant decline in
parallel with the rapid growth of the world population. Current methods
of energy consumption are considered environmentally hazardous and
contribute to global warming. To address this rapidly growing concern,
development of improved extraction techniques for renewable fuel
alternatives is underway. To date, most bioethanol is derived from
conventional food and animal feed crops such as corn and sugarcane. A
mass shift towards second generation lignocellulosic biomass-derived
biofuels will rely on techniques that will enable easy access to plant
cell wall components. Lignocellulosic conversion to biofuel requires
pretreatment with heat and acid to break the cell wall for effective
lignin release. Transgenic plants have been designed for such processes
to express reduced or modified lignin for facilitation of
saccharification. Plants have been successfully engineered to express
enhanced levels of glycoside hydrolases to enable efficient and
cost-effective polysaccharicle hydrolysis. Furthermore, strategies
focusing on increased accessibility of cellulose-degrading enzymes to
their substrate have been developed, and rely on cell wall-targeted
expression of glycoside hydrolases, cellulose binding modules (CBMs) or
other cell wall modifying proteins. A novel approach to cell wall
modification comprising the introduction of non-crystalline, soluble
polysaccharides to cell walls is discussed. Such approaches will further
advance efforts toward establishment of applicable and effective biofuel
production methods. (C) 2009 Elsevier Ireland Ltd. 

Document Number: 95

Engineering oilseed crops for industrial purposes requires a suitable
crop that does not outcross to any food oilseed crop, thus eliminating
problems of gene flow. Crambe abyssinica is such a dedicated crop as it
does not hybridize with any existing food oilseed crops. However, lack
of regeneration and transformation protocols has limited the use of C.
abyssinica in genetic manipulation studies. In this study, efficient
regeneration and transformation protocols for Crambe have been
developed. Hypocotyls of C. abyssinica cv. Galactica were incubated on a
Murashige and Skoog medium supplemented with various plant growth
regulators (PGRs). Among the different PGR combinations tested, 10 mu M
thidiazuron and 2.7 mu M alpha-naphthaleneacetic acid promoted highest
frequency of regeneration, up to 60%. Among six Agrobacterium stains
evaluated, each harbouring the cloning vector containing the neomycin
phosphotransferase (nptII) and beta-glucuronidase (gus) genes. EHA101
and AGL-1 yielded the highest transformation frequencies of 1.3 and
2.1%, respectively. Putative transgenic lines were recovered, and
confirmed as transgenic by Southern blot analysis. Subsequently,
Agrobacterium-mediated transformation of hypocotyls of cv. Galactica
with constructs harbouring the wax synthase and fatty acid reductase
genes have also successfully recovered confirmed transgenic plants
carrying these transgenes.

Document Number: 96

Plant carotenoids are C40 isoprenoids with multiple biological roles.
Breeding for carotenoid content in rice, maize and wheat is a relevant
issue, both for their importance in human health and nutrition and for
their influence in food colouration in products such as pasta from durum
wheat. Regarding human health, vitamin A deficiency (VAD) is one of the
major causes of malnutrition in the world. As many as 500 000 children
become blind due to VAD each year with many of them dying from
VAD-related illness within 1 year.
This review presents the main results in the improvement of endosperm
carotenoid levels in rice, maize and wheat considering the methodology
used, either transgenic or non-transgenic; the breeding target, such as
provitamin A or total carotenoid content; the identification of new
carotenogenic genes/alleles related to the available variation for this
trait; and the development of new functional markers for marker-assisted
selection. A comparative overview among these species and key areas for
further improvement are also identified. Carotenoid enhancement in
grasses would benefit from comparative studies among Triticeae species
since they allow the understanding of the diversity basis. Therefore,
the comparative overview given in this work will be relevant not only to
rice, maize and wheat but also to other Triticeae species.

Document Number: 97

P>Rust fungi cause devastating diseases on many important food crops,
with a damaging stem rust epidemic currently affecting wheat production
in Africa and the Middle East. These parasitic fungi propagate
exclusively on plants, precluding the use of many biotechnological tools
available for other culturable fungi. In particular the lack of a stable
transformation system has been an impediment to the genetic manipulation
required for molecular analysis of rust pathogenicity. We have developed
an Agrobacterium-mediated genetic transformation procedure for the model
flax rust fungus Melampsora lini, which infects flax (Linum
usitatissimum). Selection of transgenic rust lines is based on silencing
of AvrL567, which encodes a rust effector protein that is recognised by
the flax L6 immune receptor. The non-transgenic rust line is unable to
infect flax plants expressing L6, while silenced transgenic lines are
virulent on these plants, providing an effective selection system. This
directly confirms that the cloned AvrL567 gene is responsible for flax
rust virulence phenotypes, and demonstrates the utility of this system
to probe rust gene function.

Document Number: 98

Rapeseed (Brassica napus) is one of the most important edible oilseed
crops in the world and is increasingly used globally to produce
bio-diesel. Therefore, increasing oil content of oilseed corps is of
importance economically in both food and oil industries. The wri1 genes
are differentially expressed in B. napus lines with different oil
content. To investigate the effects of B. napus WRI1 (BnWRI1) on oil
content, two Bnwri1 genes with different lengths, Bnwri1-1 and Bnwri1-2,
were identified and sequenced. Homology analysis shows 80% amino acids
of Bnwri1s are homologous to Arabidopsis thaliana WRI1 (AtWRI1).
Overexpression of Bnwri1 cDNAs driven by cauliflower mosaic virus
35S-promoter in 51 transgenic A. thaliana lines resulted in 10-40%
increased seed oil content and enlarged seed size and mass. Detailed
analysis on transgenic embryos indicates an increased cell size other
than cell number. In addition, Bnwri1 sequence polymorphism is highly
related to oil content (p < 0.001). Taking together, Bnwri1 has
potential applications in food and oil industries and in rapeseed
breeding. (C) 2009 Elsevier Masson SAS. 

Document Number: 99

B. Schnettler, A. Gonzalez, R. Avila, H. Miranda, J. Sepulveda, and M.
Denegri. 2010. Preference for oils with different types of genetic
modifications in Temuco, Araucania Region, Chile. Cien. Inv. Agr.
37(1):17-28. Considering that attitudes towards biotechnology depend on
the type of modification used, a study was performed to determine the
importance of the type of Genetic Modification (GM) in preferences for
oil purchased by consumers in Temuco (Araucania Region, Chile) and the
existence of different market segments using a survey of 400 people. A
conjoint analysis demonstrated that the type of GM (70.3%) was more
important than the price of the oil (29.7%). Four segments were
distinguished using an analysis of hierarchical conglomerates, but the
type of GM was the most important factor in all segments. Two groups
preferred oil without GM and oil with GM that reduces the use of
pesticides and rejected oil with GM that increases calcium content; the
larger of these groups (42.8%) preferred to pay a lower price, while the
other (16.8%) they preferred higher priced oils. One group (27.2%)
rejected all types of GM. The smallest group (13.2%) rejected oil
without GM and preferred oil with GM that reduces the use of pesticides
and oil with GM that increases calcium. All of the groups rejected oil
with GM that reduces production costs. The groups were differentiated by
the family size, lifestyle, ethnic origin, reading of the labels on
foodstuffs, and the variety of oil habitually consumed. The absence of
GM in oils is desirable for the majority of consumers (86.8%); however,
a high proportion (72.8%) would accept oils with GM that reduces the use
of pesticides.

Document Number: 100

Given the important role that starch plays in food and non-food uses of
many crops, particularly wheat, efforts are being made to manipulate its
composition through modification of the amylose/amylopectin ratio.
Approaches used to achieve this goal include the manipulation of the
genes involved in the starch biosynthetic pathway using natural or
induced mutations and transgenic methods. The use of mutagenesis to
produce novel allelic variation represents a powerful tool to increase
genetic diversity and this approach seems particularly appropriate for
starch synthase genes for which limited variation exists. In this work,
an EMS-mutagenised population of bread wheat cv. Cadenza has been
screened by combining SDS-PAGE analysis of granule bound starch proteins
with a TILLING (Targeting Induced Local Lesions IN Genomes) approach at
the gene level. In particular we have focused on two groups of synthase
genes, those encoding the starch synthase II (Sgp-1) and those
corresponding to the waxy proteins (Wx). SDS-PAGE analysis of granule
bound proteins allowed the identification of single null genotypes
associated with each of the three homoeologous loci. Molecular
characterization of induced mutants has been performed using genome
specific primer pairs for Sgp-1 and Wx genes. Additional novel allelic
variation has also been detected at the different Sgp-1 homoeoloci by
using a reverse genetic approach (TILLING). In particular single
nucleotide substitutions, introducing a premature stop codon and
creating amino acid substitutions, have been identified.

Document Number: 101

Genetic engineering is becoming an important tool for the improvement of
plants for various forms of production. As varieties are developed for
both food and non-food use different production lines must be kept
separate. For good management practices of different lines an
understanding of gene-flow is essential. Barley has been proposed to be
an ideal plant species for genetic engineering as it has a low frequency
of cross-fertilization and limited seed dispersal. In the present study,
pollen-mediated exchange of genetic material between non-transgenic
closed-flowering barley variants was examined in experimental plots
under sub-arctic conditions in Iceland. The pollen-mediated dispersal
was studied using the barley varieties Golden Promise and Ven, as pollen
donor and acceptor, respectively. Only two hybrid plants were identified
from a total of 700,000 screened plants giving a hybridization frequency
of 0.0003%. It is concluded that adequate isolation distances and good
management practices should suffice to prevent cross-fertilization
between different lines of barley.

Document Number: 102

Objective
The aim of this work was to investigate the occurrence of Roundup Ready
soybean in enteral nutrition formulas sold in Brazil.
Methods
A duplex Polymerase Chain Reaction based on the amplification of the
lectin gene and the construction of the recombinant deoxyribonucleic
acid of transgenic glyphosate-tolerant soybean (35S promoter and
chloroplast transit peptide gene) was performed in order to analyze the
deoxyribonucleic acid obtained from nine soy protein isolate-containing
formulas.
Results
Despite the highly processed nature of the food matrices, amplifiable
deoxyribonucleic acid templates were obtained from all tested samples,
as judged by the amplification of the lectin gene sequence. However,
amplicons relative to the presence of Roundup Ready soybean were
restricted to one of the nine enteral nutrition formulas analyzed as
well as to the soybean reference powder, as expected. Quantitative
analysis of the genetically modified formula by real-time Polymerase
Chain Reaction showed a content of approximately 0.3% (w/w) of
recombinant deoxyribonucleic acid from the Roundup Ready soybean.
Conclusion
The results show that one of the formulas contained genetically modified
soy, pointing to the need of regulating the use of transgenic substances
and of specific labeling in this product category.

Document Number: 103

Experimental animal models are necessary to evaluate food functionality
and drug effectiveness on human disease. In vitro experiments using
effective enzymes and primary cultured cells or established cell lines
are often utilized as first screening method for some functional assay.
However final in vivo evaluation for the food function and drug action
using animal model is essential. In this review, some spontaneous
experimental animal models for human disease, stimulus-induced
experimental models in rodents, some mutation model animal or gene
transgenic animal model and gene knockout model animals are picked LIP
and presented about the outlines. Especially experimental animal models
about life-style related disease for human are described showing some
references. On the other hand, we are looking for alternative method for
experimental animals from general opinion about protection of all nature
animals. It is expected that the new alternative evaluation methods for
function of foods and drugs using new gene-technology and
cyto-technology without experimental animals are developed in near
future. (Accepted Sep. 4, 2009)

Document Number: 104

Vitamins are widely applied to produce food (including dietary
supplements), pharmaceuticals, feed-stuffs, and, also, as components of
cosmetics. On the industrial scale, the majority of vitamins are
produced using methods of chemical synthesis or through the extraction
of natural substances, but, in many cases, those processes consume high
amounts of energy and generate high waste disposal and waste utilization
costs. Those arguments were the spur for searching for options to
replace syntheses with biotechnological processes beginning from the use
of micro-organisms in the selected bio-transformations (vitamin C) to
the complete microbiological synthesis with engineered strains, for
example in the case of vitamin B12. An alternative is the production of
raw materials of plants with an increased content of vitamins by the
metabolic design of pathways of their biosynthesis, or using them as
bio-reactors, the so called `phytopharming' (vitamins A and E). This
paper presents some selected aspects relating to the biotechnological
production of vitamins and to the selection of transgenic organisms for
their production.

Document Number: 105

The Triticeae crops comprise wheat, barley, rye and triticale, which
together provide a major portion of the world's food and feed. The
growing demand for human nutrition and renewable energy requires an
intensification in application-oriented research and the establishment
and utilization of current biotechnology in these crops. Genetic
transformation provides an important means both to elucidate gene
function, and to engineer crop plants in a directed and precise way.
This review covers a range Of issues surrounding the production of
stable transgenic lines within the Triticeae. Some quality aspects of
transgenesis of particular relevance to the Triticeae are also
discussed.

Document Number: 106

In recent years, the increasing knowledge on the molecular mechanisms
underlying mineral uptake, transport, homeostasis and deposition within
plants, has paved the way for a more targeted approach to improving the
nutrient status of crop plants based on biotechnology. In the present
paper we will briefly review existing knowledge on the distribution and
transport pathways of iron in the two small grained cereals, barley and
wheat, and focus on the efforts made to increase the iron content in
cereals in general. However, mineral content is not the only factor of
relevance for improving the nutritional status of poor populations. It
is thus well documented that a number of plant components can act either
as promoters or inhibitors of mineral uptake in the human digestive
system (Frossard et al. J Sci Food Agric 80, 817-879 2000;
Brinch-Pedersen et al. J Cereal Sci 46, 308-326 2007). The nutritional
impact of increasing mineral content accordingly has to be seen in the
context of mineral bioavailability. Finally, we will briefly report on
recent data from barley, where laser capture microdissection of the
different grain tissues combined with gene expression profiling has
provided some insight into metal transport and deposition (Tauris et al.
2009). In the present paper we will provide a tentative and preliminary
roadmap for iron trafficking in the barley grain.

Document Number: 107

The earth is rapidly changing through processes such as rising [CO(2)],
[O(3)], and increased food demand. By 2050 the projected atmospheric
[CO(2)] and ground level [O(3)] will be 50% and 20% higher than today.
To meet future agricultural demand, amplified by an increasing
population and economic progress in developing countries. crop yields
will have to increase by at least 50% by the middle of the century. FACE
(Free Air Concentration Enrichment) experiments have been conducted for
more than 20 years in various parts of world to estimate. under the most
realistic agricultural conditions possible, the impact of the CO(2)
levels projected for the middle of this century on crops. The
stimulations of crop seed yields by the projected CO(2) levels across
FACE studies are about 18% on average and up to similar to 30% for the
hybrid rice varieties and vary among crops, cultivars, nitrogen levels
and soil moisture The observed increase in crop yields under the
projected CO(2) levels fall short of what would be required to meet the
projected future food demand, even with the most responsive varieties.
Crop biomass production and seed yield is the product of photosynthetic
solar energy conversion. Improvement in photosynthetic radiation use
efficiency stands as the most promising opportunity allowing for major
increases in crop yield in a future that portends major changes in
climate and crop growing environments. Our advanced understanding of the
photosynthetic process along with rapidly advancing capabilities in
functional genomics, genetic transformation and synthetic biology
promises new opportunities for crop improvement by greater
photosynthesis and crop yield Traits and genes that show promise for
improving photosynthesis are briefly reviewed, including enhancing leaf
photosynthesis capacity and reducing photorespiratron loss, manipulating
plant hormones' responses for better ideotypes, extend: rig duration of
photosynthesis, and increasing carbon partitioning to the sink to
alleviate feedback inhibition of photosynthesis (C) 2009 Elsevier
Ireland Ltd All rights reserved

Document Number: 108

There is a duality in plant tolerance to pollutants and its response to
the pollutants' stress.
On the one hand some plants, (hyper)tolerant to heavy metals, are able
to hyperaccumulate these metals in shoots, which could be beneficial for
phytoremediation purposes to clean-up soil and water. On the other hand
tolerant food crops, exposed to heavy metals in their growth medium, may
be dangerous as carriers of toxic metals in the food chain leading to
food toxicity. There is an additional duality in plant tolerance to
heavy metals and that is in food crops that are tolerant and/or
hyperaccumulators, which could be used on one hand for phytoremediation,
under controlled conditions and on the other hand for food fortification
with essential metals.
Similarly, plants are also exposed to a large number of xenobiotic
organic pollutants. Because they generally cannot avoid these compounds,
plants take up, translocate, metabolize and detoxify many of them. There
is a large variability in tolerance (defence) mechanisms against organic
pollutants among plant species. This includes production of reductants
but also scavenger molecules like ascorbate and glutathione and
expression of the P-450 defence system, and superfamilies of the enzymes
glutathione- and glucosyl-transferases. Again, with view to organic
pollutants, plant detoxification mechanisms might well protect the plant
itself, but produce compounds with some deleterious potential for other
organisms.
In this review we discuss these dualities on the basis of examples of
agricultural and 'wild' species exposed to metal contaminants (mainly
Cd) and organic pollutants. Differences in uptake and translocation of
various pollutants and their consequences will be considered. We will
separately outline the effects of the organic and non-organic Pollutants
on the internal metabolism and the detoxification mechanisms and try to
indicate the differences between both types of pollutants. Finally the
consequences and solutions of these dualities in plant tolerance to
pollutants will be discussed. (C) 2009 Elsevier B.V. All rights
reserved.

Document Number: 109

Salt stress imposes a major environmental threat to agriculture and its
adverse impacts are getting more serious problem in regions where saline
water is used for irrigation. Therefore, the efforts to increase salt
tolerance of crop plants bear remarkable importance to supply
sustainable agriculture on marginal lands and could potentially improve
crop yield overall. Acclimation of plants to salinized conditions depend
upon activation of cascades of molecular networks involved in stress
sensing, signal transduction and the expression of specific
stress-related genes and metabolites. Adaptational processes are
elaborate and more than one gene might be expressed during the
acclimation process. Isolation of Salt Overly Sensitive (SOS) genes by
sos mutants shed us light on the relationship between ion homeostasis
and salinity tolerance. The essential role of antioxidative system to
maintain a balance between the overproduction of Reactive Oxygen Species
(ROS) and their scavenging to keep them at signaling level for
reinstating metabolic homeostasis has already been established.
Compatible osmolytes synthesized to maintain equal water potential with
the environment under salinity conditions implements another strategy to
develop resistance against salinity. With the growing body of
information about molecular markers, genomics and post-genomics and thus
increasing understanding of signaling pathways and mechanisms that
contributes to plant stress responses, significant breakthroughs have
been emerged to figure out the mechanism and control of salinity
tolerance at molecular level. Many transgenic works were carried out to
produce transgenic plants to develop enhanced tolerance to salt stress.
However, a few of them seem succeeded to be implemented in salt-affected
marginal lands efficiently. This minireview focuses on the recent
developments in salinity tolerance research aiming to contribute
sustainable food production under salt stress in the face of a globally
warming ecosystem. (C) 2009 Elsevier B.V. All rights reserved

Document Number: 110

Over the years, cyanobacteria have been regarded as ideal model systems
for studying fundamental biochemical processes like oxygenic
photosynthesis and carbon and nitrogen assimilation. Additionally, they
have been used as human foods, sources for vitamins, proteins, fine
chemicals, and bioactive compounds. Aiming to increase plant
productivity as well as nutritional values, cyanobacterial genes
involved in carbon metabolism, fatty acid biosynthesis, and pigment
biosynthesis have been intensively exploited as alternatives to
homologous gene sources. In this short review, transgenic plants with
cyanobacterial genes generated over the last two decades are examined,
and the future prospects for transgenic crops using cyanobacterial genes
obtained from functional genomics studies of numerous cyanobacterial
genomes information are discussed.

Document Number: 111

P>Nearly one-third of the world's population, mostly women and children,
suffer from iron malnutrition and its consequences, such as anaemia or
impaired mental development. Iron fortification of food is difficult
because soluble iron is either unstable or unpalatable, and non-soluble
iron is not bioavailable. Genetic engineering of crop plants to increase
iron content has therefore emerged as an alternative for iron
biofortification. To date, strategies to increase iron content have
relied on single genes, with limited success. Our work focuses on rice
as a model plant, because it feeds one-half of the world's population,
including the majority of the iron-malnourished population. Using the
targeted expression of two transgenes, nicotianamine synthase and
ferritin, we increased the iron content of rice endosperm by more than
six-fold. Analysis of transgenic rice lines confirmed that, in
combination, they provide a synergistic effect on iron uptake and
storage. Laser ablation-inductively coupled plasma-mass spectrometry
showed that the iron in the endosperm of the transgenic rice lines
accumulated in spots, most probably as a consequence of spatially
restricted ferritin accumulation. Agronomic evaluation of the high-iron
rice lines did not reveal a yield penalty or significant changes in
trait characters, except for a tendency to earlier flowering. Overall,
we have demonstrated that rice can be engineered with a small number of
genes to achieve iron biofortification at a dietary significant level.

Document Number: 112

P>The growth of genetically engineered maize that produces the
insecticidal protein Cry3Bb1 from Bacillus thuringiensis (Bt) is an
effective method to control corn rootworms (Diabrotica spp.), which are
threatening maize production in North America and Europe. In this study,
the risk of Cry3Bb1-expressing maize for the predatory spider Theridion
impressum, a common species in European maize fields, was assessed.
Quantification of Cry3Bb1 in potential prey species collected in Bt
maize plots and prey spectrum analysis revealed that T. impressum
ingests Cry3Bb1 in the field. Exposure to the Bt protein, however, was
highly variable because some potential prey species, such as
phloem-feeding herbivores and predators, contained little or no Cry3Bb1,
whereas leaf-feeding herbivores contained high concentrations. Adult and
juvenile T. impressum spiders were fed with Cry3Bb1-containing food
(prey or maize pollen) for 8 weeks in the laboratory to examine the
toxicity of the Bt protein. No differences in mortality, weight
development or offspring production were observed between spiders
provided with food containing or not containing Cry3Bb1. Retrospective
power analysis indicated that the bioassays were sufficiently sensitive
to detect meaningful differences if present. Although Cry3Bb1 is
ingested by the spider in the field, our data provide no evidence for
toxicity. Consequently, the growth of corn rootworm-resistant Bt maize
appears to pose no risk for T. impressum.

Document Number: 113

Ferritins are major players in plant iron homeostasis. Surprisingly,
their overexpression in transgenic plants led only to a moderate
increase in seed iron content, suggesting the existence of control
checkpoints for iron loading and storage in seeds. This work reports the
identification of two of these checkpoints. First, measurement of seed
metal content during fruit development in Arabidopsis thaliana reveals a
similar dynamic of loading for Fe, Mn, Cu, and Zn. The step controlling
metal loading into the seed occurs by the regulation of transport from
the hull to the seed. Second, metal loading and ferritin abundance were
monitored in different genetic backgrounds affected in vacuolar iron
transport (AtVIT1, AtNRAMP3, AtNRAMP4) or plastid iron storage (AtFER1
to 4). This approach revealed (1) a post-translational regulation of
ferritin accumulation in seeds, and (2) that ferritin stability depends
on the balance of iron allocation between vacuoles and plastids. Thus,
the success of ferritin overexpression strategies for iron
biofortification, a promising approach to reduce iron-deficiency anemia
in developing countries, would strongly benefit from the identification
and engineering of mechanisms enabling the translocation of high amounts
of iron into seed plastids.

Document Number: 114

Cereals provide more than half the world population's calorific intake,
and have a variety of other important uses as food and beverage
ingredients, livestock feeds, and as sources of renewable energy and
industrial components. The technology to genetically modify many
important cereals is now well-established, thereby presenting new
opportunities to produce cereals with enhanced quality and novel
properties. In 2007, GM (genetically modified) maize with insect and
herbicide resistance was grown on over 30 million hectares worldwide,
yet to date, there are no GM cereals with enhanced or novel grain
(end-use) qualities being grown in commercial farmers' fields. This
review will discuss some of the latest GM technology developments
reported to enhance the quality of cereals for food and other uses.
Developments and opportunities involving gene manipulation for starch
and protein quality, as well as non-starch polysaccharides, phenolic
compounds and micronutrients will also be discussed. The current paucity
of GM cereals with enhanced grain quality is not related to the absence
of technological progress, rather it is the regulatory and consumer
acceptance issues that have slowed the release of these crops.

Document Number: 115

In recent years, special concerns have been raised about the safety
assessment of foods and food ingredients derived from genetically
modified organisms (GMOs). A growing number of countries establish
regulations and laws for GMOs in order to allow consumers an informed
choice. In this case, a lot of methods have been developed for the
detection of GMOs. However, the reproducibility among methods and
laboratories is still a problem. Consequently, it is still in great
demand for more effective methods. In comparison with the gel
electrophoresis, the capillary electrophoresis (CE) technology has some
unique advantages, such as high resolution efficiency and less time
consumption. Therefore, some CE-based methods have been developed for
the detection of GMOs in recent years. All kinds of CE detection
methods, such as ultraviolet (UV), laser induced fluorescence (LIF), and
chemiluminescence (CL) detection, have been used for GMOs detection.
Microchip capillary electrophoresis (MCE) methods have also been used
for GMOs detection and they have shown some unique advantages.

Document Number: 116

Sufficient methionine levels in the seed are critical for the supply of
a balanced diet for feed and food. Currently, animal feed is
supplemented with chemically synthesized methionine, which could be
completely replaced with naturally synthesized methionine. However,
insufficient levels of methionine are due to alleles of two genes in the
maize genome that are expressed during seed development, which have a
high percentage of methionine codons, ranging from 23 to 28%, while free
methionine is very low. The two genes, dzs10 and dzs18, belong to the
prolamin gene family that arose during the evolution of the grasses and
were duplicated during a whole genome duplication event. We have found
several dzs10 and dzs18 null alleles caused either by transposon
insertion or frame shift mutations. Maize seeds with null mutations of
both genes have a normal phenotype in contrast to other prolamin genes,
explaining the accumulation of methionine deficiency in normal breeding
efforts. Moreover, the trans-regulation of these genes deviates from
Mendelian inheritance. One allele of the regulatory locus dzr1 is
inherited in a parent-of-origin fashion, while another allele appears to
prevent Mendelian segregation of the high-methionine phenotype in
backcrosses.

Document Number: 117

Genetic engineering of chloroplasts normally requires the stable
introduction of bacterial derived antibiotic or herbicide-resistance
genes as selective markers. Ecological and health concerns have been
raised due to the presence of such genes within the environment or the
food supply. One way to overcome this issue is the use of plant genes
able to confer a metabolic or developmental advantage to the transformed
cells manipulating the plant's biosynthetic pathways. We explored the
feasibility of using, for plastid transformation, the selection system
based on the feedback-insensitive anthranilate synthase (AS)
alpha-subunit gene of tobacco (ASA2) as a new selective marker and the
indole analogue 4-methylindole (4MI) or the tryptophan analogue
7-methyl-DL-tryptophan (7MT) as the selection agents. An expression
cassette containing Prrn-ASA2 was effectively integrated into the region
between accD and ycf4 of the tobacco plastome by the biolistic process.
Plastid transgenic plants were obtained on medium supplemented with 300
mu M 7MT or 4MI. Transplastomic plants showed normal phenotype and
fertility and the resistance to the selection agents 7MT and 4MI was
transmitted maternally. The plastid transformed lines also exhibited a
higher level of AS enzyme activity that was less sensitive to
Trp-feedback inhibition and, consequently, increased free Trp levels in
leaves about 7-fold.

Document Number: 118

From the origins the man has looked and selected vegetables with
nutritive value for larger productivity. The knowledge of DNA structure
allowed genetic engineering to develop and supplying tools for the
accomplishment of specific alterations in the genome considerably. The
products of these alterations are denominated transgenic or organisms
genetically modified (OGM) and they present high application potential
in several areas of the human activity as: agriculture, medicine,
health, production and processing of foods, biochemical production and
control of diseases. Nowadays, transgenic plants, originating from
technology of the DNA recombinant, brought new varieties cultivated
already in more than 100 million hectares in 23 countries, including
Brazil, where 8 varieties were already approved for the National
Technical Commission of Biosafety (CTNBio). That method of genetic
improvement facilitated the introduction of desirable characteristics in
plants, such as, resistance to biotic and non-biotic stress and
optimization of the composition of some essential nutrients to the
animal and human health. While these progresses of the biotechnology
open new perspectives for the solution of problems in areas as the
agriculture, the transgenic liberation for use in the nature brings
concerns as for possible problems of ecological nature and for the human
and animal health. These concerns brought the creation of government
agencies to control the use of this technology and to regulate the
safety of transgenic foods and yours derived. Until the moment, the
scientific studies show that the transgenic liberated commercially are
so safe or more to the environment and the animal and human health that
the conventional ones.

Document Number: 119

Resveratrol, a plant phenolic compound, is found in grapes and red wine,
but is not widely distributed in other common food sources. The pathway
for resveratrol biosynthesis is well characterized. Metabolic
engineering of this compound has been achieved in tomato plants
(Lycopersicon esculentum Mill.) in order to improve their nutritional
value. Tomato plants synthesizing resveratrol were obtained via the
heterologous expression of a grape (Vitis vinifera L.) cDNA encoding for
the enzyme stilbene synthase (StSy), under the control of the
fruit-specific promoter TomLoxB. The resulting LoxS transgenic plants
accumulated trans-resveratrol and trans-piceid, in particular in the
skin of the mature fruits. Quantitative analyses carried out on LoxS
fruits were compared with those of a tomato line constitutively
expressing the stsy gene (35SS). The LoxS fruits contained levels of
trans-resveratrol that were 20-fold lower than those previously reported
for the 35SS line. The total antioxidant capability and ascorbate
content in transformed fruits were also evaluated, and a significant
increase in both was found in the LoxS and 35SS lines. These results
could explain the higher capability of transgenic fruits to counteract
the pro-inflammatory effects of phorbol ester in monocyte-macrophages
via the inhibition of induced cyclo-oxygenase-2 enzyme.

Document Number: 120

With the ever increasing number of genetically modified plants
authorized worldwide, including in the European Union, high throughput
detection methods need to be developed. In this paper, a
quadruplex-real-time-PCR method is described which allows rapid and
simultaneous screening of food for the presence of target DNA sequences
from the cauliflower mosaic virus 35S promoter, the NOS terminator from
Agrobacterium tumefaciens, the soya reference lectin gene and the maize
reference alcool dehydrogenase gene (adh). Three of the four primers and
probe combinations have already been published elsewhere, whereas
primers and probe for NOS terminator-PCR were developed in-house.
Validation data show sensitivities down to five copies for 35S promoter
and NOS terminator PCR, even when target sequences of the competing PCRs
are in large excess. Thorough adjustment of primer and probe
concentrations allowed high individual PCR efficiencies with negligible
physical cross-talk between the four detection channels. This method
provides a basis for a rapid screening of food for the most frequently
used regulatory elements present in GM crops authorized for food in the
European Union. In addition it provides information about the presence
of species which are possibly genetically modified.

Document Number: 121

In this paper I critique two popular, non-scientific attitudes toward
genetically engineered foods. In doing so, I will be employing the
concepts of ambiguity, purity/impurity, control/resistance, and
unity/diversity as developed by Latina feminist metaphysicians. I begin
by casting a critical eye toward a specific anti-biotech account of
transgenic food crops, an account that I will argue relies on an
anti-feminist metaphysics. I then cast that same critical eye toward a
specific pro-biotech account, arguing that it also relies on such an
anti-feminist metaphysics. I will argue further that this metaphysics
yields a less accurate account of genetics. I end by arguing that if we
adopt a Latina feminist metaphysics we can more accurately understand
plants, genetics, and genetic engineering.

Document Number: 122

Meeting the requirement for absorbed iron is difficult for vegetarians,
and their iron status often is lower than that of nonvegetarians. Beans
contain ferritin in low concentrations, but it is possible to enhance
this content by plant breeding or by inserting the gene for ferritin
into plants, eg, soybeans. Because each ferritin molecule can bind to
thousands of iron atoms, this may be a sustainable means to increase the
iron contents of plants. Before such efforts are launched, it is
important to determine whether iron in ferritin is bioavailable. This
has been assessed in vitro by using human intestinal (Caco-2) cells and
in vivo by using radiolabeled ferritin and whole-body counting in human
subjects. Dietary factors affecting iron absorption, eg, ascorbic acid,
phytate, and calcium, had limited effect on iron uptake from intact
ferritin by Caco-2 cells, which suggests that ferritin-bound iron is
absorbed via a mechanism different from that of nonheme iron. In an in
vitro digestion system, ferritin was shown to be relatively resistant to
proteolytic enzymes. Binding of ferritin to Caco-2 cells was shown to be
saturable, and the kinetics for binding were characteristic of a
receptor-mediated process. In human subjects, iron from purified soybean
ferritin given in a meal was as well absorbed as iron from ferrous
sulfate. In conclusion, iron is well absorbed from ferritin and may
represent a means of biofortification of staple foods such as soybeans.
Am J Clin Nutr 2009; 89(suppl): 1680S-5S.

Document Number: 123

Substantial agricultural production increases are necessary in order to
feed the growing world population. This also requires harnessing the
potentials of modern biotechnology, complementing more traditional crop
improvement techniques. Insect-resistant transgenic plants have already
proved to deliver higher effective yields through better crop
protection. However, hunger is not only a production but also a
distribution problem, meaning that many people are simply too poor to
have sufficient access to food. Transgenic crops can play a role in this
respect too, as large parts of the world's poor are dependent on
agriculture as a source of income. For Bt cotton in particular, effects
on poverty reduction have already been observed in some developing
countries. But not every transgenic application is suitable for small
scale farmers, and the social impacts also depend on the institutional
environment. Therefore, technological approaches should always be seen
as a component of a broader development strategy. Rationalizing the
emotional biotechnology debate would be desirable.

Document Number: 124

The wheat grain is the most important organ for human food and therefore
is the target for much research focused on modifying its composition to
improve nutritional and functional components. Genetic transformation
provides a precise tool to alter the composition of wheat grain by
expressing new genes or by down-regulating groups of proteins encoded by
multigene families such as gliadins, which contain clusters of epitopes
that are active groups in triggering celiac disease. For such work,
specific promoters are required to express such constructs in the wheat
endosperm. In the present study we report the isolation and
characterization of a gamma-gliadin promoter from transgenic wheat, and
the analysis of gliadin synthesis during grain development in bread
wheat by Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass
Spectrometry (MALDI/TOF MS). The gamma-gliadin promoter fragment was
isolated from bread wheat by genome walking and was re-introduced,
driving the expression of the gusA gene, by particle bombardment, giving
fifteen independent transgenic lines. Detailed analysis of the sequence
of the 885 bp promoter fragment showed that it contains three prolamin
boxes but only one is conserved according to the consensus sequence
reported. The AACA/TA motif is present twice in published gamma-gliadin
promoter sequences. The RY element i.e., CATGCAT or CATGCAC, is also
present twice in the published promoter. Transgenic lines were
classified as high, medium, and low expressers. The expression of the
gusA gene was found only in the seeds of the transgenic lines. GUS
staining was first detected in the outer endosperm of the lobes, and
then it extended to the whole outer endosperm. GUS staining was not
found in the aleurone layer nor in the embryo. The qRT-PCR data
confirmed the data obtained by GUS staining. The expression of the gusA
gene determined by qRT-PCR for the high expresser line (B281) was 4 and
8 times higher than that of medium (B282) and low (B286) expresser
lines, respectively. MALDI/TOF-MS showed that gliadins exhibited
different patterns of synthesis during the course of seed maturation.
Thus, gliadins with masses higher than 36,000 Da were synthesised within
the first 12 days post anthesis while those with masses lower than
36,000 Da were synthesised later. Results of GUS staining, qRT-PCR and
MALDI/TOF-MS showed that the gamma-gliadin promoter reported in this
work could be a good candidate to downregulate wheat gliadins.

Document Number: 125

Immunoassays are used to screen for the presence of genetically modified
organisms in raw materials. However, processing may condition the
usefulness of immunoassays to analyse genetically modified foods because
it leads to protein denaturation that affects recognition by antibodies.
We studied the effect of processing on the detection of Cry1A(b) protein
in model foods prepared with transgenic maize using a sandwich ELISA.
Nixtamalization at 100 A degrees C for 5 min and at 85 A degrees C for
60 min gave 40 and 70% loss of Cry1A(b) protein. In the preparation of
porridge, the concentration of Cry1A(b) protein did not change until the
mixture reached 75 A degrees C, but it decreased by 90% after 3 min at
that temperature. Concentration of Cry1A(b) protein decreased by 90% in
tortillas griddled at 180 A degrees C for 20 s, but no protein was
detected in fried tortillas after 10 s at 190 A degrees C. Cry1A(b)
protein is rapidly denatured by heat treatment resulting in a marked
decline in concentration and decreased detection in processed foods.

Document Number: 126

Work with cereals (barley and wheat) and a legume (Medicago truncatula)
has established thioredoxin h (Trx h) as a central regulatory protein of
seeds. Trx h acts by reducing disulfide (S-S) groups of diverse seed
proteins (storage proteins, enzymes, and enzyme inhibitors), thereby
facilitating germination. Early in vitro protein studies were
complemented with experiments in which barley seeds with Trx h
overexpressed in the endosperm showed accelerated germination and early
or enhanced expression of associated enzymes (alpha-amylase and
pullulanase). The current study extends the transgenic work to wheat.
Two approaches were followed to alter the expression of Trx h genes in
the endosperm: (1) a hordein promoter and its protein body targeting
sequence led to overexpression of Trx h5, and (2) an antisense construct
of Trx h9 resulted in cytosolic underexpression of that gene
(Arabidopsis designation). Underexpression of Trx h9 led to effects
opposite to those observed for overexpression Trx h5 in
barley-025EFretardation of germination and delayed or reduced expression
of associated enzymes. Similar enzyme changes were observed in
developing seeds. The wheat lines with underexpressed Trx showed delayed
preharvest sprouting when grown in the greenhouse or field without a
decrease in final yield. Wheat with overexpressed Trx h5 showed changes
commensurate with earlier in vitro work: increased solubility of
disulfide proteins and lower allergenicity of the gliadin fraction. The
results are further evidence that the level of Trx h in cereal endosperm
determines fundamental properties as well as potential applications of
the seed.

Document Number: 127

Nixtamalised flour from transgenic maize (genetically modified maize
with the cDNA of amarantin) and its tortillas were evaluated for some
technological and nutritional properties and compared to the commercial
brand MASECA(TM). Nixtamalised transgenic maize flour (NTMF) showed
higher protein content, total colour difference, pH, water solubility
index, essential amino acids content and lower Hunter "L" value, water
absorption index, resistant starch and retrograded resistant starch than
MASECA(TM) flour. Tortillas from NTMF had higher protein content (12.64%
vs 8.93%, db), essential amino acids content and calculated protein
efficiency ratio (C-PER: 2.05 vs 1.04) than tortillas from MASECA(TM).
Tortillas from both nixtamalised transgenic maize and MASECA(TM) flours
showed similar sensory properties (puffing and acceptability). The use
of transgenic maize for flour and tortilla preparation may have a
positive impact on the nutritional status of people from countries where
maize is the basic staple food. (C) 2008 Elsevier Ltd. All rights
reserved.

Document Number: 128

The characterisation of transgenic and non-transgenic soybeans was
performed in this work based on the examination of their protein
profiles obtained by rapid chromatographic techniques. Two
reversed-phase chromatographic methods using monolithic and perfusion
stationary phases were applied to the separation of soybean proteins
from different transgenic and non-transgenic soybeans. The development
of the monolithic LC methodology was carried out through the study of
the influence of different parameters such as gradient, ion-pairing
reagent, and temperature on the separation of soybean proteins. Results
from monolithic LC analysis were compared with those obtained by
perfusion LC using a method previously developed by our research team.
Perfusion and monolithic LC enabled the separation of soybean proteins
in less than 3 and 8 min, respectively. In both cases, there were
certain features in the chromatograms that seemed to be characteristic
of transgenic samples. A deeper analysis of chromatographic profiles was
then performed by the application of multivariate classification
techniques. Results from these multivariate techniques showed that the
two methods presented similar classification capabilities being both
Suitable for the characterisation of transgenic and non-transgenic
soybeans. A more robust mathematical model was built With the data
obtained by the perfusion method using 10 additional samples for
training (a total of 26 samples), obtaining a 96.2% of correct
classification. This model was validated by a cross-validation procedure
(80.8% of correct classification) and by the correct classification of
15 Out of 16 blind samples. (C) 2008 Elsevier Ltd. 

Document Number: 129

A growth performance experiment was conducted to assess the feeding
value of a double-stacked transgenic corn grain for growing-finishing
pigs. The genetically modified corn grain contained event DAS59122-7,
which expresses the Cry34/35Ab1 binary insecticidal protein for the
control of corn rootworm. This modified transgenic grain is resistant to
western corn rootworm and is also tolerant to herbicides containing the
active ingredient glufosinate-ammonium. The modified grain (59122), a
nontransgenic near-isoline grain (control corn), and a commercial corn
(Pioneer brand hybrid 35P12) were grown in a 2005 production trial in
individually isolated plots that were located 201 m apart. A total of
108 pigs were allotted to corn-soybean meal diets containing 1 of the 3
grains as the sole source of corn. There were 3 pigs per pen and 12
replicate pens per treatment. Pigs were fed grower diets from 37 to 60
kg, early finisher diets from 60 to 90 kg, and late finisher diets from
90 to 127 kg. Within each phase, data for ADG, ADFI, and G: F were
calculated. At the conclusion of the experiment, pigs were slaughtered
and data for carcass quality were collected. Differences between 59122
and the control corn were evaluated, with statistical significance at P
< 0.05. No differences in ADG, ADFI, or G: F between pigs fed the
control corn and pigs fed the modified corn were observed during the
grower, early finisher, or late finisher phases. For the entire
experimental period, no difference between pigs fed the control and the
59122 corn were observed for final BW (128.9 vs. 127.1 kg), ADG (1.02
vs. 1.00 kg), ADFI (2.88 vs. 2.80 kg), or G: F (0.356 vs. 0.345 kg/kg).
Likewise, no differences in dressing percentage (76.48 vs. 76.30%), LM
area (49.8 vs. 50.4 cm(2)), 10th-rib back fat (2.20 vs. 2.12 cm), and
carcass lean content (52.9 vs. 53.4%) were observed between pigs fed the
control and the 59122 corn grain. It was concluded that the nutritional
value of the modified transgenic corn grain containing event DAS-59122-7
was similar to that of the nontransgenic near-isoline control.

Document Number: 130

Uptake and translocation of micronutrients are essential for plant
growth. These micronutrients are also important food components. We
generated transgenic rice plants over-expressing OsIRT1 to evaluate its
functional roles in metal homeostasis. Those plants showed enhanced
tolerance to iron deficiency at the seedling stage. In paddy fields,
this over-expression caused plant architecture to be altered. In
addition, those plants were sensitive to excess Zn and Cd, indicating
that OsIRT1 also transports those metals. As expected, iron and zinc
contents were elevated in the shoots, roots and mature seeds of
over-expressing plants. This demonstrates that OsIRT1 can be used for
enhancing micronutrient levels in rice grains.

Document Number: 131

Oilseed rape (Brassica napus L.) is an important agronomic crop used for
oil, food and animal feed. Glyphosate resistant B. napus is tolerant to
the herbicide glyphosate and has been widely adopted due to higher
economic returns. However, a concern for growing transgenic B. napus is
the potential escape of cp4 epsps transgene via cross fertilization with
wild and/or weedy species including Brassica juncea, especially in
China, where B. juncea is prevalent. We assessed transgene inheritance
and quantification upon wild B. juncea x transgenic B. napus outcrossing
from three backcross (BC) generations and their progenies using
real-time PCR. Under in vitro conditions, a stable inheritance and
similar cp4 epsps copy number between different generations (BC2, BC3
and BC4) and their progenies were observed. A marginally higher copy
number was observed when B. juncea was used as the paternal as compared
to when it was used as maternal parent. This study demonstrates stable
inheritance of the glyphosate resistance trait and the production of fit
out crossed offspring in cases of wild type B. juncea x transgenic B.
napus.

Document Number: 132

BACKGROUND: Rice is one of the most important staple food crops for more
than half the global population, who rely on it for as much as 80% of
their diet. By one estimate, the world population is projected to grow
to approximately 11 billion people by the year 2050. So it is a
formidable task to meet the future demand. For this reason, breeders
make a great deal of effort to produce new rice varieties with traits
such as higher yield, improved nutritional content and better resistance
to disease and pests, via transgenic biotechnological protocols. Dozens
of transgenic rice lines have been developed since the first transgenic
rice plant production in the late 1980s. With the rapid approach of
transgenic rice commercialisation, it is becoming necessary to develop
techniques capable of detecting and quantifying genetically modified
(GM) rice.
RESULT: Here we describe a method in which transgenic DNA is quantified
by amplifying part of the 35S-CaMV promoter and standardising it against
an amplified portion of an endogenous single copy, rice specific gene
encoding sucrose phosphate synthase. Both reactions are performed
simultaneously in a single tube. Standard calibration curves were
developed by diluting DNA extracted from a blend of non-transgenic
(c.v., Nipponbare) and 5% KMD2 transgenic rice. The method was tested
for the quantification of the five GM rice events, including KMD2, Wan
21A, GC-1, H1597 and TR4, which contain the 35S-CaMV promoter. The
coefficient of variation varied from 3.15% to 12.84%, which is up to
acceptance criterion over the dynamic range of the method.
CONCLUSION: In this study, we successfully applied a multiplex real-time
PCR assay to GM rice, which employed SPS as the endogenous reference
gene and the gene regulation element 35S-CaMV promoter as a GMO marker.
The detection limit and limit of quantification is sufficient to comply
with all relevant regulations in the EU and worldwide. The detection
system could be applied in routine analysis for the quantification of GM
rice in food materials, such as instant rice, unpolished rice, rice
flours, biscuit powders, and starch. It may prove useful with regard to
a robust screening technique of broad utility as transgenic rice enters
global commodity markets. (C) 2009 Society of Chemical Industry

Document Number: 133

Regions traditionally destined to agriculture report an ever increasing
exposure to cold and drought conditions. This is especially important in
countries like Chile where crop management options are limited. The
development of new cultivars with better yields under adverse conditions
is fundamental if the ever increasing demand for food is to be matched;
however, improving tolerance to abiotic stresses has proved to be a
complex task. In this regard, development in plant physiology and
genomics in the last 20 years has led to a deeper understanding of how
plants respond to stress and mechanisms responsible for different ranges
of tolerance observed in nature. This review discusses the techniques
currently most in use in gene expression analysis, together with some
important experimental design variables, such as the developmental stage
of the plant, stress intensity and duration, and how different stresses
may interact when performing assays. On the other hand, it is
fundamental to properly select gene expression techniques according to
the available information on the genome, the crop and the final
objective of the research. All these points must be considered to ease
transition from genomics to practical applications to crop species in
order to increase their tolerance to stress. In this regard, the rapid
development of new techniques in gene expression analysis with lower
costs will determine a new revolution in crop research in coming
decades. Therefore, Chile needs to be prepared in this area to continue
its development as a major food producer worldwide.

Document Number: 134

The development of transgenic plants may help alleviate both
environmental and food problems. In Japan, transgenic plants cannot be
planted in the field without undergoing an environmental biosafety
assessment under confined and semi-confined conditions. The main
objectives of environmental biosafety assessment studies are to define
the properties of the host plant and to evaluate its influence on other
organisms. To appraise the influence of a transgenic plant on other
plants, the transgenic plant is examined for new compounds that
influence plant using the sandwich method to assay the allelopathic
activity of leachate from dried leaf samples. However, because
Eucalyptus leaves are difficult to dry, the sandwich method does not
allow the evaluation of multiple samples at the same time. Here, we
report a new "homogenized method" that relies on homogenizing fleshy
leaf samples, instead of drying them. The alleopathic activity of a
non-transgenic plant and an Antisense-lim transgenic plant were
evaluated using both the sandwich method and the homogenized method to
determine whether the homogenized method was available for in biosafety
for in biosafety assessments of transgenic plants. The homogenized
method may be an effective and useful tool for evaluating differences
between non-transgenic and transgenic plants using multiple, concurrent
samples.

Document Number: 135

The storage of fruits and vegetables at low temperature near the
freezing point is the foremost technology applied to retard postharvest
ripening and to extend the shelf-life period of agricultural products.
However, most tropical and subtropical produce is sensitive to chilling
injury, which constitutes a set of physiological alterations caused by
exposure to low temperatures for variable time periods, to the detriment
of quality. This article is a thorough review of the physiological,
hormonal and molecular mechanisms involved in the induction and
development of this physiopathy. Also, the different postharvest
technologies of a chemical, physical or biotechnological nature assayed
In research or applied in the agro-food industry with the aim of
inhibiting or delaying the emergence of chilling injury in sensitive
plant produce of agricultural interest are reviewed. (C) 2009 Society of
Chemical Industry

Document Number: 136

BACKGROUND: The Cry toxins are already a useful alternative or
supplement to synthetic chemical pesticide application in commercial
agriculture and forest management.
RESULTS:The Cry1ab/ac gene from Bacillus thuringiensis was cloned from
the genome of genetically modified rice by polymerase chain reaction
(PCR). Owing to the large number of Escherichia coli low-usage codons in
the Cry1ab/ac gene, the first 20 codons were optimised by PCR to improve
the expression of the Cry1ab/ac gene in E. coli. The Cry1Ab/Ac protein
was highly expressed in E. coli as inclusion bodies that could be
dissolved in 8 mol L(-1) urea and purified on a His Trap (TM) crude
column under denaturing conditions. The purified Cry1Ab/Ac protein was
dialysed in refolding buffers to obtain a soluble and biologically
active protein. To achieve better biological activity, the His-tag was
digested from the Cry1Ab/Ac protein with enterokinase, and the Cry1Ab/Ac
protein was further purified by gel filtration on a fast performance
liquid chromatography Superclex 75 HR 10/30 column using an AKTA
purifier. The identity of the purified Cry1Ab/Ac protein sequence was
confirmed by western blot and matrix-assisted laser desorption
ionisation time-of-flight mass spectrometry. The final purified
Cry1Ab/Ac protein was 99.2% pure and retained its biological activity,
as determined in a growth inhibition assay of Chilo suppressalis.
CONCLUSION: The purified Cry1Ab/Ac protein could be used to evaluate the
food safety of transgenic plants containing the Cry1ab/ac gene and to
produce antibodies for immune-based methods employed in the detection of
genetically modified organisms containing the Cry1ab or Cry1ac gene. It
might also serve as a new biological insecticide to reduce the use of
broad-spectrum insecticides. (C) 2009 Society of Chemical Industry

Document Number: 137

The efficacy of the ipt-type Multi-Auto-Transformation (MAT) vector
system to transform the extensively grown cassava cultivar "KU50" was
evaluated. This system utilizes the isopentenyltransferase (ipt) gene as
morphological marker for visual selection of transgenic lines. The
extreme shooty phenotype (ESP) of transgenic lines is lost due to the
removal of ipt gene mediated by the yeast Rint/RS system. As a result,
phenotypically normal shoots, considered marker-free transgenic plants,
could be obtained. When transforming KU50 cassava cultivar with two
different ipt-type MAT vectors, transformation frequency at 19-21% was
observed. Among the total number of ESP explants, 32-38% regained normal
extended shoot phenotype and 88-96% of which were confirmed to represent
the marker-free transgenic plants. This is the first demonstration of
the efficacy of Rint/RS system in promoting excision of ipt marker gene
in cassava specie, with the consequent rapid production of marker-free
transgenic plants. The high efficiency of this system should facilitate
pyramiding a number of transgenes by repeated transformation without
having to undergo through laborious, expensive and time-consuming
processes of sexual crossing and seed production. The generation of
marker-free, thus environmentally safe, genetically modified cassava
clones should also ease the public concerns regarding the use of
transgenic cassava in both food and nonfood industries.

Document Number: 138

Transgenic cows secreting over 3 mu g of lysostaphin/mL of milk are
protected against mastitis caused by Staphylococcus aureus, but it is
unknown if active lysostaphin persists through dairy processing
procedures or affects the production of fermented dairy foods. The
objective of this study was to determine the fate of lysostaphin as milk
was pasteurized and then processed into cheese. Raw milk from transgenic
cows was heat treated at 63 degrees C for 30 min, 72 degrees C for 15 s
(high temperature, short time), or 140 C for 2 s (UHT). Portions of the
high temperature, short-time milk were manufactured into semi-hard
cheeses. Aliquots taken at each processing step were assayed to
determine the quantity (ELISA) and activity (ability to inhibit S.
aureus growth) of lysostaphin. Results indicated that most of the
lysostaphin was present in the aqueous portion of the milk and was not
affected by pasteurization, although UHT treatment reduced enzyme
concentration by 60%. The quantity and activity of the lysostaphin
decreased during cheesemaking. Based on the amount of lysostaphin
present in the starting cheesemilk, 10 to 15% of the lysostaphin was
recovered in the whey, 21 to 55% in the cheese curd at d 1, and 21 to
36% in cheese stored at 4 C for 90 d. Enough of the lysostaphin secreted
into milk by transgenic cows survived typical dairy processing
conditions to impart potential value as a bioprotective agent against
staphylococci in dairy foods.

Document Number: 139

Dioxin residues widely contaminate soil and agricultural products at low
concentrations and may accumulate in organisms at the top of food chains
owing to their physicochemical properties. In this study, we have
developed novel, dioxin-inducible, reporter gene expression systems
regulated by recombinant aryl hydrocarbon receptors (AhRs). The
recombinant AhRs, referred to as XDVs, consist of the DNA-binding domain
of the bacterial repressor protein LexA, a 90-kDa heat shock protein-
and ligand-binding regulatory domain from mouse AhR, and the
transactivation domain of herpes simplex virus regulatory protein VP16.
Transgenic tobacco plants carrying XDVs absorb various AhR ligands,
including 3-methylcholanthrene, beta-naphthoflavone and indigo from
solid medium and vermiculite, and show dose- and time-dependent
expression of the beta-glucuronidase reporter gene. The results clearly
suggest that XDVs are functional transcription factors that respond to
AhR ligands, and that the XDV-mediated reporter gene expression system
is applicable to bioassays for dioxin residues in the environment.

Document Number: 140

Small interfering RNAs (siRNAs) and microRNAs (miRNAs) are effector
molecules of RNA interference (RNAi), a highly conserved RNA-based gene
suppression mechanism in plants, mammals and other eukaryotes.
Endogenous RNAi-based gene suppression has been harnessed naturally and
through conventional breeding to achieve desired plant phenotypes. The
present study demonstrates that endogenous small RNAs, such as siRNAs
and miRNAs, are abundant in soybean seeds, corn kernels, and rice grain,
plant tissues that are traditionally used for food and feed. Numerous
enclogenous plant small RNAs were found to have perfect complementarity
to human genes as well as those of other mammals. The abundance of
enclogenous small RNA molecules in grain from safely consumed food and
feed crops such as soybean, corn, and rice and the homology of a number
of these dietary small RNAs to human and animal genomes and
transcriptomes establishes a history of safe consumption for dietary
small RNAs. (C) 2008 Elsevier Ltd. 

Document Number: 141

Abscisic Acid (ABA) is an important phytohormone involved in abiotic
stress resistance in plants. A group of bZIP transcription factors play
important roles in the ABA signaling pathway in Arabidopsis. However,
little is known about the function of their orthologs in rice, where
they may hold a great potential for developing drought resistant food
crops. In this study, our phylogenetic analysis showed that this group
of bZIPs was evolutionarily conserved between Arabidopsis and rice,
which implies that they may share similar functions. We demonstrated
with quantitative RT-PCR that the expressions of most of these OsbZIPs
were significantly induced by ABA, ACC, and abiotic stresses. OsbZIP72,
a member of this group, was proved to be an ABRE binding factor in rice
using the yeast hybrid systems. We showed that it could bind to ABRE and
transactivate the downstream reporter genes in yeast, and the
transactivity was depending on its N-terminal region. Transgenic rice
overexpressing OsbZIP72 showed a hypersensitivity to ABA, elevated
levels of expression of ABA response gene such as LEAs, and an enhanced
ability of drought tolerance. These results suggest that OsbZIP72 plays
a positive role in drought resistance through ABA signaling, and is
potential useful for engineering drought tolerant rice.

Document Number: 142

A method was developed using enhanced liquid chromatography coupled with
electrospray ionization mass spectrometry for the analysis and
quantitation of 2 main potato glycoalkaloids, alpha-chaconine, and
alpha-solanine, without any pre-concentration or derivatisation steps.
Calibration curves generated by this technique exhibited a linear
dynamic range from 0.025 to 50 mu g/mL and from 0.05 to 50 mu g/mL for
alpha-chaconine and alpha-solanine, respectively. Matrix effects were
evaluated by comparing calibration curves measured in matrix-matched and
solvent-based systems. Ion suppression due to matrix effects was weak
and extraction recoveries of 88 to 114% were obtained in different
sample matrices spiked with analyte concentrations ranging from 15 to 35
mu g/mL. Potatoes that had been genetically modified to tolerate
glufosinate contained the same glycoalkaloid levels as their
non-transgenic counterpart. We suggest complementing compositional
comparison assessment strategy by validating quantitative analytical
methods for the toxic glycoalkaloids in potato plants.

Document Number: 143

The addition of soybean proteins to processed meat products has
significantly increased in recent years due to the interesting
functional and nutritional properties of these vegetable proteins. Since
the Roundup Ready (RR) soybean is the only transgenic soybean line
approved for market in EU this work was aimed at monitoring its presence
in meat products on the Serbian food market. The extracted DNA was
analyzed using duplex polymerase chain reaction (PCR) with primer pairs
aimed at the lectin gene and 35S promoter. Samples positive for the
presence of GM soybean were subjected to a real-time quantification of
the percentage of RR soya. The results indicated that out of fifty
processed meat products examined, twelve gave positive results with 35S
promoter and all contained RR soya below 0.1%. (C) 2008 Elsevier Ltd.

Document Number: 144

Cassava is a valuable source of food for developing countries, but it
contains highly toxic cyanogen compounds and antinutrients. Cyanogens
are found in 3 forms in cassava: cyanogenic glucoside (95% linamarin and
5% lotaustratin), cyanohydrins, and free cyanide. Different processing
techniques exist to remove cyanogens and their effectiveness depends on
the processing steps and the sequence utilized, and it often is
time-dependent. Pounding or crushing is the most effective for
cyanogenic glucoside removal because it ruptures cell compartments, thus
allowing direct contact between linamarin and the enzyme linamarase that
catalyzes the hydrolytic breakdown. Crushing and sun-drying cassava
roots made into flour removes 96% to 99% of total cyanogens, whereas
soaking and sun-drying into lafun or fufu, or soaking and fermenting and
roasting into gari or farina, removes about 98% of cyanogens. For
cassava leaves, which have 10 times more cyanogens than roots, pounding
and boiling in water is an efficient process to remove about 99% of
cyanogens. Other strategies to reduce toxicity include development of
low-cyanogen cassava varieties and cassava transgenic lines with
accelerated cyanogenesis during processing. Although phytate and
polyphenols have antioxidant properties, they interfere with digestion
and uptake of nutrients. Fermentation and oven-drying are efficient
processing methods to remove phytate (85.6%) and polyphenols (52%),
respectively, from cassava roots. Sun-drying the leaves, with or without
prior steaming or shredding, removes about 60% phytate. Cassava is a
nutritionally strategic famine crop for developing countries and,
therefore, reducing its toxicity and improving its nutritional value is
crucial.

Document Number: 145

The risk assessment of genetically modified (GM) crops for human
nutrition and health has not been systematic. Evaluations for each GM
crop or trait have been conducted using different feeding periods,
animal models, and parameters. The most common result is that GM and
conventional sources induce similar nutritional performance and growth
in animals. However, adverse microscopic and molecular effects of some
GM foods in different organs or tissues have been reported. Diversity
among the methods and results of the risk assessments reflects the
complexity of the subject. While there are currently no standardized
methods to evaluate the safety of GM foods, attempts towards
harmonization are on the way. More scientific effort is necessary in
order to build confidence in the evaluation and acceptance of GM foods.

Document Number: 146

Genetic engineering provides a means to introduce genes into plants via
mechanisms that arc different in some respects from classical breeding.
A number of commercialized, genetically engineered (GE) varieties, most
notably canola, cotton, maize and soybean, were created using this
technology, and at present the traits introduced are herbicide and/or
pest tolerance. In 2007 these GE crops were planted in developed and
developing countries on more than 280 million acres (113 million
hectares) worldwide, representing nearly 10% of rainfed cropland.
Although the United States leads die world in acres planted with GE
crops, the majority of this planting is on large acreage farms. In
developing countries, adopters are mostly small and resource-poor
farmers. For farmers and many consumers worldwide, planting and eating
GE crops and products made from them are acceptable and even welcomed;
for others GE crops raise food and environmental safety questions, as
well as economic and social issues. In Part I of this review, some
general and food issues related to GE crops and foods were discussed. In
Part II, issues related to certain environmental and socioeconomic
aspects of GE crops and foods are addressed, with responses linked to
the scientific literature.

Document Number: 147

The alpha ' and beta subunits of soybean beta-conglycinin were expressed
in rice seeds in order to improve the nutritional and physiological
properties of rice as a food. The alpha ' subunit accumulated in rice
seeds at a higher level than the beta subunit, but no detectable
difference in mRNA transcription level between subunits was observed.
Sequential extraction results indicate that the alpha ' subunit formed
one or more disulphide bonds with glutelin. Electron microscopic
analysis showed that the alpha ' subunit and the beta subunit were
transported to PB-II together with glutelin. In mature transgenic seeds,
the P subunit accumulated in low electron density regions in the
periphery of PB-II, whereas the alpha ' subunit accumulated together
with glutelin in high-density regions of the periphery. The subcellular
localization of mutated alpha ' subunits lacking one cysteine residue in
the N-terminal mature region (alpha ' Delta Cys1) or five cysteine
residues in the pro and N-terminal mature regions (alpha ' Delta Cys5)
were also examined. Low-density regions were formed in PB-II in mature
seeds of transgenic rice expressing alpha ' Delta Cys 5 and alpha '
Delta Cys1. alpha ' Delta Cys5 was localized only in the low-density
regions, whereas alpha ' Delta Cys1 was found in both low- and
high-density regions. These results suggest that the alpha ' subunit
could make a complex via one or more disulphide bonds with glutelin and
accumulate together in PB-II of transgenic rice seeds.

Document Number: 148

As genetically modified (GM) foods are starting to intrude in our diet
concerns have been expressed regarding GM food safety. These concerns as
well as the limitations of the procedures followed in the evaluation of
their safety are presented. Animal toxicity studies with certain GM
foods have shown that they may toxically affect several organs and
systems. The review of these studies should not be conducted separately
for each GM food, but according to the effects exerted on certain organs
it may help us create a better picture of the possible health effects on
human beings. The results of most studies with GM foods indicate that
they may cause some common toxic effects such as hepatic, pancreatic,
renal, or reproductive effects and may alter the hematological,
biochemical, and immunologic parameters. However, many years of research
with animals and clinical trials are required for this assessment. The
use of recombinant GH or its expression in animals should be re-examined
since it has been shown that it increases IGF-1 which may promote
cancer.

Document Number: 149

Plant-based foods offer an array of nutrients that are essential for
human nutrition and promote good health. However, the major staple crops
of the world are often deficient in some of these nutrients. Traditional
agricultural approaches can marginally enhance the nutritional value of
some foods, but the advances in molecular biology are rapidly being
exploited to engineer crops with enhanced key nutrients. Nutritional
targets include elevated mineral content, improved fatty acid
composition, increased amino acid levels, and heightened antioxidant
levels. Unfortunately, in many cases the benefits of these
"biofortified" crops to human nutrition have not been demonstrated.

Document Number: 150

Genetically modified ( GM) crops are now grown commercially in 23
countries, with another 29 granting approval for import and release into
the environment. Despite the socio-economic and environmental benefits
of the technology, further development is being hampered by differences
in national regulatory frameworks relating to research, biosafety, and
to the trade and use of GM crops. The biosafety regulations in different
countries are based on five main international instruments that
influence the development of national biosafety systems in terms of
field trial permit requirements, risk assessment criteria, labeling,
traceability, transparency, public awareness, post-monitoring and import
regulations. The global harmonization of data collection, testing
procedures and information exchange would help to remove artificial
trade barriers, expedite the adoption of GM crops, foster technology
transfer and protect developing countries from exploitation, instilling
confidence and bringing the benefits of GM products to the consumer.

Document Number: 151

Soybean is a major crop species providing valuable feedstock for food,
feed and biofuel. In recent years, considerable progress has been made
in developing genomic resources for soybean, including on-going efforts
to sequence the genome. These efforts have identified a large number of
soybean genes, most with unknown function. Therefore, a major research
priority is determining the function of these genes, especially those
involved in agronomic performance and seed traits. One means to study
gene function is through mutagenesis and the study of the resulting
phenotypes. Transposon-tagging has been used successfully in both model
and crop plants to support studies of gene function. In this report, we
describe efforts to generate a transposon-based mutant collection of
soybean. The Ds transposon system was used to create activation-tagging,
gene and enhancer trap elements. Currently, the repository houses
approximately 900 soybean events, with flanking sequence data derived
from 200 of these events. Analysis of the insertions revealed
approximately 70% disrupted known genes, with the majority matching
sequences derived from either Glycine max or Medicago truncatula
sequences. Among the mutants generated, one resulted in male-sterility
and was shown to disrupt the strictosidine synthase gene. This example
clearly demonstrates that it is possible to disrupt soybean gene
function by insertional mutagenesis and to derive useful mutants by this
approach in spite of the tetraploid nature of the soybean genome.

Document Number: 152

The use of viral vectors is a method for introducing foreign genes into
various animal species. Vectors based on retro-, adeno-, flavi-, and
parvoviruses have been used for research in animal species of
agricultural importance, such as chickens, quail, swine, cows, goats,
sheep, fish, crustaceans, and mollusks. Viral vectors allow for
efficient transgenic integration into host genome or for transient
expression of the transgenic construct in somatic tissues. Because of
that, viral vectors are important tools for research and potentially
other biotechnology applications such as improving animal production
qualities and introducing disease resistance, thus improving food
quality and safety. Other uses may include generating animal models of
human diseases and using animals as bioreactors for production of
therapeutic proteins. Each vector type provides a unique set of
advantages and limitations, which are in some cases specific to an
animal species or a method of introduction. This article discusses viral
vector characteristics and potential applications in agriculturally
important animal species. It discusses advantages and disadvantages of
using viral vectors in genetic engineering of agricultural animals.

Document Number: 153

Does transgenically incorporated insect resistance affect constitutive
and herbivore-inducible terpenoid emissions and multitrophic
communication under elevated atmospheric CO(2) or ozone (O(3))? This
study aimed to clarify the possible interactions between allocation to
direct defences Bacillus thuringiensis (Bt) toxin production) and that
to endogenous indirect defences under future climatic conditions.
Terpenoid emissions were measured from vegetative-stage non-Bt and Bt
Brassica napus grown in growth chambers under control or doubled CO(2),
and control (filtered air) or 100 ppb O(3). The olfactometric
orientation of Cotesia vestalis, an endoparasitoid of the herbivorous
diamondback moth (Plutella xylostella), was assessed under the
corresponding CO(2) and O(3) concentrations.
The response of terpenoid emission to CO(2) or O(3) elevations was
equivalent for Bt and non-Bt plants, but lower target herbivory reduced
herbivore-inducible emissions from Bt plants. Elevated CO(2) increased
emissions of most terpenoids, whereas O(3) reduced total terpenoid
emissions. Cotesia vestalis orientated to host-damaged plants
independent of plant type or CO(2) concentration. Under elevated O(3),
host-damaged non-Bt plants attracted 75% of the parasitoids, but only
36.8% of parasitoids orientated to host-damaged Bt plants.
Elevated O(3) has the potential to perturb specialized food-web
communication in Bt crops.

Document Number: 154

The photosynthetic performance of two transgenic Medicago truncatula
lines engineered for water deficit (WD) resistance and a non-transformed
line was assessed in a growth chamber experiment in well-watered, WD and
stress recovery conditions. Direct gas exchange measurements showed that
the transgenic plants had lower photosynthetic rates under well-hydrated
conditions when compared to the non-transformed line. Photosynthesis
light curves confirmed this difference but more importantly showed a
progressive change in photosynthetic behaviour with intensity of
dehydration. Dehydration led to sharp decreases of maximum
photosynthesis (A(max)), photosynthetic apparent quantum yield () and
apparent light compensation point. The recovery rates showed that all
plant lines had a similar capacity to regain control photosynthetic
values. Furthermore, results suggested that light was more limiting for
photosynthesis than atmospheric CO(2) concentration. The results are
discussed in terms of the use of photosynthesis light response curves as
a non-destructive and expeditious approach to select M. truncatula
transformants with improved WD resistance.

Document Number: 155

The herbicide-tolerant transgenic rapeseed Topas 19/2 (synonym HCN92)
has been approved for environmental release in Canada, Japan, Australia
and the USA, and exported to a number of other countries as raw
material. The purpose of this study was to establish event-specific
qualitative and quantitative detection methods for Topas 19/2. The
T-integration junction sequence spanning the host plant DNA and the
integrated transgene of the Topas 19/2 event was isolated and
identified. The event-specific qualitative detection method was
established to produce an amplicon of 110 basepairs (bp) with an
absolute detection limit of 10 initial template copies. The
event-specific quantitative detection method was developed with the
limit of detection (LOD) and limit of quantification (LOQ) being
approximately 5 and 50 initial template copies, respectively. The
developed real-time PCR systems were assessed using two mixed rapeseed
samples with known Topas 19/2 contents. Expected results were obtained.
Crown Copyright (c) 2008 Published by Elsevier Ltd. 

Document Number: 156

Plant-made pharmaceuticals (PMPs) offer promise as efficient and
cost-effective products for the treatment Of human and animal diseases.
An advantage of producing pharmaceuticals in maize is the large storage
capacity and stability for proteins and starch in seed, allowing for
manufacturing recombinant proteins Such as antigens and antibodies.
Other advantages of the maize system include safety, high yields, and
scalability of production and processing. However, the benefits of this
technology Must be balanced against potential health and environmental
risks that may be associated with its use. Because PMPs presently have
no provision for regulatory tolerance, their inadvertent occurrence in
foods and feeds remains an important economic consideration, even when
the health and environmental risks are low. Pollen drift is considered.
Source of potential contamination of maize-made pharmaceuticals in the
food chain. In addition to physical and temporal isolation requirements,
open field pharmaceutical maize production also calls for controlled
pollen release. Here, we describe two strategies to address the issue of
transgenic pollen drift. First, we describe the development and genetic
transformation of a tissue culture-amenable male-sterile line using
biolistic- or Agrobacterium-mediated transformation methods. Secondly,
we describe the introgression of a transgene from male-fertile
transgenic maize to male-sterile germplasm by conventional breeding.
After six seasons of breeding, this second strategy allows us to obtain
100% transgenic seeds from an open-field production using a
non-transgenic line as the pollinator.

Document Number: 157

Food matrices are controlled for the presence of authorised and
unauthorised GMOs (Genetically Modified Organisms). Transgenic papaya
was developed and approved for the market in the USA. However,
authorisation in EC have not been accomplished. Recently, unauthorised
GM papaya was catch on EC market. Therefore it is necessary to dispose
by an assay revealing genetically modified fruits. High quality DNA is
required for the assay. We tested and optimised CTAB, Wizard and spin
column based procedure for DNA extraction. We found that CTAB based
protocol allows isolation of amplifiable DNA from fresh fruit flesh and
stones, whereas spin columns alowed isolation of amplifiable DNA from
candied fruits. Modified protocol is described. No transgenic fresh
fruit or candied fruit from the market was detected.

Document Number: 158

Present method allows a simple and rapid identification of maize and
specific transgenic soyabean event in raw materials and processed feed
samples. The method combines two aspects: suitable DNA purification and
amplification by means of multiplex RT-PCR. Efficiency and accuracy of
this method have been tested and the limit of detection (LOD) reached
was of 0.1 ng/mu l for one of the species and of 1 ng/mu l for the other
one. The checking of the method has been realized by means of RT-PCR
using the specific probes for each one of the systems by means of
comparison with certified reference materials. The obtained results
showed that with this method it is possible to obtain, in a shorter
time, quality values for both: identification and quantification of
GMOs, in raw and processed samples.

Document Number: 159

Transgene expression from the plant's plastid genome represents a
promising strategy in molecular farming because of the plastid's
potential to accumulate foreign proteins to high levels and the
increased biosafety provided by the maternal mode of organelle
inheritance. In this article, we explore the potential of transplastomic
plants to produce human immunodeficiency virus (HIV) antigens as
potential components of an acquired immunodeficiency syndrome (AIDS)
vaccine. It is shown that the HIV antigens p24 (the major target of
T-cell-mediated immune responses in HIV-positive individuals) and Nef
can be expressed to high levels in plastids of tobacco, a non-food crop,
and tomato, a food crop with an edible fruit. Optimized p24-Nef fusion
gene cassettes trigger antigen protein accumulation to up to
approximately 40% of the plant's total protein, demonstrating the great
potential of transgenic plastids to produce AIDS vaccine components at
low cost and high yield.

Document Number: 160

A major limitation of crop biotechnology and breeding is the lack of
efficient molecular technologies for precise engineering of target
genomic loci. While transformation procedures have become routine for a
growing number of plant species, the random introduction of complex
transgenenic DNA into the plant genome by current methods generates
unpredictable effects on both transgene and homologous native gene
expression. The risk of transgene transfer into related plant species
and consumers is another concern associated with the conventional
transformation technologies. Various approaches to avoid or eliminate
undesirable transgenes, most notably selectable marker genes used in
plant transformation, have recently been developed. These approaches
include cotransformation with two independent T-DNAs or plasmid DNAs
followed by their subsequent segregation, transposon-mediated DNA
elimination, and most recently, attempts to replace bacterial T-DNA
borders and selectable marker genes with functional equivalents of plant
origin. The use of site-specific recombination to remove undesired DNA
from the plant genome and concomitantly, via excision-mediated DNA
rearrangement, switch-activate by choice transgenes of agronomical, food
or feed quality traits provides a versatile "transgene maintenance and
control" strategy that can significantly contribute to the transfer of
transgenic laboratory developments into farming practice. This review
focuses on recent reports demonstrating the elimination of undesirable
transgenes (essentially selectable marker and recombinase genes) from
the plant genome and concomitant activation of a silent transgene (e.g.,
a reporter gene) mediated by different site-specific recombinases driven
by constitutive or chemically, environmentally or developmentally
regulated promoters. These reports indicate major progress in excision
strategies which extends application of the technology from annual,
sexually propagated plants towards perennial, woody and vegetatively
propagated plants. Current trends and future prospects for optimization
of excision-activation machinery and its practical implementation for
the generation of transgenic plants and plant products free of undesired
genes are discussed.

Document Number: 161

Genetically modified (GM) papaya lines have been approved for
commercialization and are widely cultivated in many countries. As a step
towards the development of reliable qualitative and quantitative PCR
methods for detecting GM papayas, one papaya (Fructus Caricae Carica
papaya L.) species specific gene, papain, was selected and validated as
suitable for using as an endogenous reference gene in transgenic papaya
PCR detection. In this article, both qualitative and quantitative PCR
assays for the papain gene were assayed with 11 different papaya
varieties and identical amplification products were obtained with all of
them. No amplified fragments could be detected when DNA samples from 18
kinds of other species were used as templates, which demonstrated that
this system was specific for papaya. In real-time quantitative PCR
analysis, the detection limit was as low as 10 pg of DNA. Southern blot
analysis confirmed that the papain gene was two copies in the tested
papaya varieties and no allelic variation was testified among these
tested papaya varieties. In addition, the common used exogenous genes of
the coat protein (CP) and the replicase (RP) were also assayed in
qualitative and real-time quantitative PCR.

Document Number: 162

The International Maize and Wheat Improvement Center (CIMMYT) aims to
genetically enhance both crops and generate public sector-provided
products for the resource poor, e.g., drought tolerant wheat and insect
resistant maize, and through international-national partnerships
facilitate the acquisition of improved germplasm for non-mandate crops
in the cropping systems where maize and wheat thrives; e.g., GM-papaya
through a national food security undertaking in Bangladesh. The Center
also engages in public awareness campaigns in projects such as Insect
Resistance Maize for Africa (IRMA), which includes food, feed and
environmental safety, monitoring of resistance and establishment of
refugia, non-target effects and gene flow. Monitoring of genetic
resources is a wide concern among the centers of the Consultative Group
on International Agricultural Research (CGIAR), with an emphasis on the
quality of gene banks. Decisions, policies and procedures about
monitoring should be science-based, and this requires education, an area
where CIMMYT and other CGIAR centers can play an important role. There
will be a need to continue to evaluate the need for, and type of
monitoring, as new (and unique) products are developed and released in
the emergent economies of the world.

Document Number: 163

Considering the high level of concern caused by genetically modified
foods (GMF) in developed countries, the relevance of this variable in
decision-making about the purchase of fluid milk among consumers in
Temuco (La Araucania Region, Chile) was determined. By means of a
personal survey of 400 people and using conjoint analysis, it was
determined that the presence of genetic modi. cations in food was more
important (44.7%) than brand (29.5%) and price (25.5%) in the
decision-making process. By cluster analysis three segments were
identified; the largest group (46.5%) gave similar relevance to food
production and brand, preferring genetically modified milk. For the
second group (41.5%), the presence or absence of genetic modi. cation
was the most important variable in the purchase, with a strong rejection
of milk produced by genetic manipulation and preference for
non-transgenic milk. The minority segment (12.0%) placed a higher value
on price. These three segments prefer national brand milk, reject the
product with private brands and react positively to lower prices. The
most sensitive segment to genetic food manipulation had a higher
proportion of people under 35 years of age and with no children. It is
concluded that the absence of genetic manipulation in food is a
desirable condition, mainly for young consumers of La Araucan a Region.

Document Number: 164

The increasing presence of products derived from genetically modified
(GM) plants in human and animal diets has led to the development of
detection methods to distinguish biotechnology-derived foods from
conventional ones. The conventional and real-time PCR have been used,
respectively, to detect and quantify GM residues in highly processed
foods. DNA extraction is a critical step during the analysis process.
Some factors such as DNA degradation, matrix effects, and the presence
of PCR inhibitors imply that a detection or quantification limit,
established for a given method, is restricted to a matrix used during
validation and cannot be projected to any other matrix outside the scope
of the method. In Brazil, sausage samples were the main class of
processed products in which Roundup Ready (R) (RR) soybean residues were
detected. Thus, the validation of methodologies for the detection and
quantification of those residues is absolutely necessary. Sausage
samples were submitted to two different methods of DNA extraction:
modified Wizard and the CTAB method. The yield and quality were compared
for both methods. DNA samples were analyzed by conventional and
real-time PCR for the detection and quantification of Roundup Ready (R)
soybean in the samples. At least 200 ng of total sausage DNA was
necessary for a reliable quantification. Reactions containing DNA
amounts below this value led to large variations on the expected GM
percentage value. In conventional PCR, the detection limit varied from
1.0 to 500 ng, depending on the GM soybean content in the sample. The
precision, performance, and linearity were relatively high indicating
that the method used for analysis was satisfactory.

Document Number: 165

Many industrial products and functional foods can be obtained from cheap
and renewable raw agricultural materials. For example, starch can be
converted to bioethanol as biofuel to reduce the current demand for
petroleum or fossil fuel energy. On the other hand, starch can also be
converted to useful functional ingredients, such as high fructose and
high maltose syrups, wine, glucose, and trehalose. The conversion
process involves fermentation by microorganisms and use of biocatalysts
such as hydrolases of the amylase superfamily. Amylases catalyze the
process of liquefaction and saccharification of starch. It is possible
to perform complete hydrolysis of starch by using the fusion product of
both linear and debranching thermostable enzymes. This will result in
saving energy otherwise needed for cooling before the next enzyme can
act on the substrate, if a sequential process is utilized. Recombinant
enzyme technology, protein engineering, and enzyme immobilization are
powerful tools available to enhance the activity of enzymes, lower the
cost of enzyme through large scale production in a heterologous host,
increase their thermostability, improve pH stability, enhance their
productivity, and hence making it competitive with the chemical
processes involved in starch hydrolysis and conversions. This review
emphasizes the potential of using biocatalysis for the production of
useful industrial products and functional foods from cheap agricultural
produce and transgenic plants. Rice was selected as a typical example to
illustrate many applications of biocatalysis in converting low-value
agricultural produce to high-value commercial food and industrial
products. The greatest advantages of using enzymes for food processing
and for industrial production of biobased products are their
environmental friendliness and consumer acceptance as being a natural
process.

Document Number: 166

The estimation of the percentage of transgenic Bt maize in maize flour
mixtures has been achieved in this work by high-performance liquid
chromatography using perfusion and monolithic columns and chemometric
analysis. Principal component analysis allowed a preliminary study of
the data structure. Then, linear discriminant analysis was used to
develop decision rules to classify samples in the established categories
(percentages of transgenic Bt maize). Finally, linear regression (LR)
and multivariate regression models (namely, principal component analysis
regression (PCR), partial least squares regression (PLS-1), and multiple
linear regression (MLR)) were assayed for the prediction of the
percentages of transgenic Bt maize present in a maize flour mixture.
Using the relative areas of the protein peaks, MLR provided the best
models and was able to predict the percentage of transgenic Bt maize in
flour mixtures with an error of +/-5.3%, +/-2.3%, and +/-3.8% in the
predictions of Aristis Bt, DKC6575, and PR33P67, respectively. (C) 2008
Elsevier Ltd. 

Document Number: 167

Genetically modified crops are widely grown in the world today. Labeling
is required when genetically modified organisms (GMOs) are placed on the
market. There is a need to establish a specific method for the detection
of genetically modified foods. MON863 transgenic maize containing a
Cry3Bb1 sequence that produces insecticidal protein cry3Bb1 is a major
GMO crop. In this paper, we report studies that designed specific PCR
primers and TaqMan probes based upon the 5'-transgene integration
sequence, and developed qualitative and quantitative PCR conditions
using these primers and probes. We determined the 5'-transgene
integration sequence using a ligation-mediated polymerase chain reaction
(LM PCR) method. In qualitative PCR studies, the limit of detection
(LOD) was 0.5% for MON863 in 100 ng genomic DNA. In the quantitative PCR
assays, the limit of detection (LOD) and limit of quantitation (LOQ) are
10 and 100 haploid copies, respectively. Maize samples with different
contents of genetically modified component were tested using the
established TaqMan real-time PCR system. (C) 2008 Elsevier Ltd. All
rights reserved.

Document Number: 168

Prevention of transgene flow from genetically modified crops to food
crops and wild relatives is of concern in agricultural biotechnology. We
used genes derived from food crops to produce complete male sterility as
a strategy for gene confinement as well as to reduce the food purity
concerns of consumers. Anther-specific promoters (A3, A6, A9, MS2, and
MS5) were isolated from Brassica oleracea and B. rapa and fused to the
beta-glucuronidase (GUS) reporter gene and candidate genes for male
sterility, including the cysteine proteases BoCysP1 and BoCP3, and
negative regulatory components of phytohormonal responses involved in
male development. These constructs were then introduced into Arabidopsis
thaliana. GUS analyses revealed that A3, A6, and A9 had tapetum-specific
promoter activity from the anther meiocyte stage. Male sterility was
confirmed in tested constructs with protease or gibberellin insensitive
(gai) genes. In particular, constructs with BoCysP1 driven by the A3 or
A9 promoter most efficiently produced plants with complete male
sterility. The tapetum and middle layer cells of anthers expressing
BoCysP1 were swollen and excessively vacuolated when observed in
transverse section. This suggests that the ectopic expression of
cysteine protease in the meiocyte stage may inhibit programmed cell
death. The gai gene also induced male sterility, although at a low
frequency. This is the first report to show that plant cysteine
proteases and gai from food crops are available as a novel tool for the
development of genetically engineered male-sterile plants.

Document Number: 169

Nutritional assessment of transgenic crops used for human food and
animal feed is an important aspect of safety evaluations. An
herbicide-tolerant rice (Bar68-1) was generated by the stable insertion
of bar gene, isolated from Streptomyces hygroscopicus strain HP632 into
the genome of a conventional variety of Chinese rice (D68). Nutrients
including proximates, amino acids, fatty acids. minerals and vitamins
and anti-nutritive components such as phytic acid, lectin, and trypsin
inhibitors of the brown and milled rice grain from the resulted Bar68-1
rice were compared with those of the parental line D68. The data
demonstrated that the nutritional quality of both the brown and milled
rice grains from the transgenic line was substantial equivalent to those
of the non-transgenic counterpart, and most results of nutritional
components fell within the range of values reported for other commercial
lines. (c) 2008 Elsevier Inc. 

Document Number: 170

Coexistence among genetically modified (GM) and non-GM cropping systems
and identity preservation at the field level are increasingly important
issues in many countries. Different types of pollen-mediated gene flow
(cross-fertilization) models have been released during the past decade,
primarily as a decision-support tool to achieve the European Union (EU)
0.9% GM adventitious presence (AP) labelling threshold for food and
feed. We review key empirical or mechanistic models for four diverse
crop types-canola or oilseed rape (OSR) (Brassica napus L.), maize (Zea
mays L.), wheat (Triticum aestivum L.), and creeping bentgrass (Agrostis
stolonifera L.). Their strengths, weaknesses, relevance, and utility in
simulating pollen-mediated gene flow are examined. Many empirical models
simulate gene flow well, although their utility is often restricted by
datasets with limited environmental variability or spatial scale. Few
mechanistic models have been developed, reflecting the challenge in
accurately simulating pollen-mediated gene flow by wind or insects: such
models have not been validated for commercial field scenarios. Many
models tend to provide upper-end or worst-case outcrossing predictions
and management recommendations, either because of experimental design
underlying datasets, biological and environmental stochasticity, or
chosen statistical analysis. Both experimental results and modelling
predictions of outcrossing in OSR, maize, and wheat reveal that
isolation distance or a pollen barrier (buffer zone) generally is only
recommended between small grain maize fields (ca. <5 ha) to maintain
field-average AP due to pollen-mediated gene flow below the EU
threshold. Recent advances in modelling pollen-mediated gene flow in
commercial fields are encouraging, but simulating gene flow in
heterogeneous landscapes remains an elusive goal. Moreover, practical,
user-friendly decision-support tools are needed to inform and guide
farmers in implementing coexistence measures. Crown Copyright (C) 2008
Published by Elsevier B.V. 

Document Number: 171

Purpose of review
In addition to extending lifespan, animal research shows that specific
diets benefit brain functioning. Indeed, it has been proven that caloric
restriction prevents age-related neuronal damage. What are those
mechanisms involved in the effects of caloric restriction on brain
functioning? Could caloric restriction be proposed in the future to
prevent or treat neurodegenerative disorders such as Alzheimer's
disease? Is there a future for caloric restriction interventions in
adults?
Recent findings
Hypotheses linking caloric restriction to cognitive capability include
anti-inflammatory mechanisms, reduction of neural oxidative stress,
promotion of synaptic plasticity, induction of various stress and
neurotrophic/neuroprotective factors. Caloric restriction may also
prevent beta-amyloid neuropathology in Alzheimer transgenic models.
Finally, both exercise and caloric restriction enhance neurogenesis via
different mechanisms suggesting that their combination may decrease the
risk of neurodegenerative disease.
Summary
It is now well established that caloric restriction could be used to
promote successful brain aging. Data from randomized controlled trials
in humans are limited. No positive effect on cognitive impairment was
found probably due to methodological limitations. The long-term effects
of caloric restriction in adults must be clarified before engaging in
such preventive strategy. Additional animal studies must be conducted in
the future to test the effects of 'multidomain' interventions (caloric
restriction plus regular exercise) on age-related cognitive decline.

Document Number: 172

In recent years, questions related to molecular composition and its
implications for nutrition and health have been raised as advances in
technology speed up the introduction of new diversity into breeding
programs, either via transgenic technology or by using molecular markers
in combination with wide crosses. Metabolite profiling offers great
opportunities for characterization of this diversity phenotypically with
respect to its metabolite composition. It provides a powerful resource
to guide breeding programs and to alert researchers to positive or
detrimental traits at an early stage. The power of this approach will be
vastly increased by combining it with transcript profiling and a
systematic survey of the metabolite composition of the plant products
that are already on the market. This integrated approach and holistic
profiling within a systems biology approach enables the careful tracking
of the response of the organism to conditional perturbations at
different molecular and genetic levels using available databases. This
approach to profiling will not only provide a baseline for comparison of
plants with novel traits (PNTs) with traditional comparators that are
'generally recognized as safe', but also provide a rational framework
for risk assessment via 'substantial equivalence'. It also provides
important inputs into nutritional research and contributes to the public
debate about the acceptability of changes in food-production chains and
development of science based regulation of plants with novel traits.

Document Number: 173

The genetic modification in fruit and vegetables could lead to changes
in metabolic pathways and, therefore, to the variation of the molecular
pattern, with particular attention to antioxidant compounds not
well-described in the literature. The aim of the present study was to
compare the quality composition of transgenic wheat (Triticum durum L.),
corn (Zea mays L.), and tomato (Lycopersicum esculentum Mill.) to the
nontransgenic control with a similar genetic background. In the first
experiment, Ofanto wheat cultivar containing the tobacco rab1 gene and
nontransgenic Ofanto were used. The second experiment compared two
transgenic lines of corn containing Bacillus thuringiensis "Cry toxin"
gene (PR33P67 and Pegaso Bt) to their nontransgenic forms. The third
experiment was conducted on transgenic tomato (Lycopersicum esculentum
Mill.) containing the Agrobacterium rhizogenes rolD gene and its
nontransgenic control (cv. Tondino). Conventional and genetically
modified crops were compared in terms of fatty acids content,
unsaponifiable fraction of antioxidants, total phenols, polyphenols,
carotenoids, vitamin C, total antioxidnt activity, and mineral
composition. No significant differences were observed for qualitative
traits analyzed in wheat and corn samples. In tomato samples, the total
antioxidant activity (TAA), measured by FRAP assay, and the naringenin
content showed a lower value in genetically modified organism (GMO)
samples (0.35 mmol of Fe2+ 100 g(-1) and 2.82 mg 100 g(-1),
respectively), in comparison to its nontransgenic control (0.41 mmol of
Fe2+ 100 g(-1) and 4.17 mg 100 g(-1), respectively). On the basis of the
principle of substantial equivalence, as articulated by the World Health
Organization, the Organization for Economic Cooperation and Development,
and the United Nations Food and Agriculture Organization, these data
support the conclusion that GM events are nutritionally similar to
conventional varieties of wheat, corn, and tomato on the market today.

Document Number: 174

Vitamin A deficiency (VAD) affects over 250 million people worldwide and
is one of the most prevalent nutritional deficiencies in developing
countries, resulting in significant socio-economic losses. Provitamin A
carotenoids such as beta-carotene, are derived from plant foods and are
a major source of vitamin A for the majority of the world's population.
Several years of intense research has resulted in the production of
'Golden Rice 2' which contains sufficiently high levels of provitamin A
carotenoids to combat VAD. In this report, the focus is on the
generation of transgenic maize with enhanced provitamin A content in
their kernels. Overexpression of the bacterial genes crtB (for phytoene
synthase) and crtI (for the four desaturation steps of the carotenoid
pathway catalysed by phytoene desaturase and zeta-carotene desaturase in
plants), under the control of a 'super gamma-zein promoter' for
endosperm-specific expression, resulted in an increase of total
carotenoids of up to 34-fold with a preferential accumulation of
beta-carotene in the maize endosperm. The levels attained approach those
estimated to have a significant impact on the nutritional status of
target populations in developing countries. The high beta-carotene trait
was found to be reproducible over at least four generations. Gene
expression analyses suggest that increased accumulation of beta-carotene
is due to an up-regulation of the endogenous lycopene beta-cylase. These
experiments set the stage for the design of transgenic approaches to
generate provitamin A-rich maize that will help alleviate VAD.

Document Number: 175

Genetic modification of the flavonoid pathway has been used to produce
novel colours and colour patterns in ornamental plants as well as to
modify the nutritional and pharmaceutical properties of food crops. It
has been suggested that co-ordinate control of multiple steps of the
pathway with the help of regulatory genes would lead to a more
predictable control of metabolic flux. Regulation of anthocyanin
biosynthesis has been studied in a common ornamental plant, Gerbera
hybrida (Asteraceae). An R2R3-type MYB factor, GMYB10, shares high
sequence similarity and is phylogenetically grouped together with
previously characterized regulators of anthocyanin pigmentation. Ectopic
expression of GMYB10 leads to strongly enhanced accumulation of
anthocyanin pigments as well as to an altered pigmentation pattern in
transgenic gerbera plants. Anthocyanin analysis indicates that GMYB10
specifically induces cyanidin biosynthesis in undifferentiated callus
and in vegetative tissues. Furthermore, in floral tissues enhanced
pelargonidin production is detected. Microarray analysis using the
gerbera 9K cDNA array revealed a highly predicted set of putative target
genes for GMYB10 including new gene family members of both early and
late biosynthetic genes of the flavonoid pathway. However, completely
new candidate targets, such as a serine carboxypeptidase-like gene as
well, as two new MYB domain factors, GMYB11 and GMYB12, whose exact
function in phenylpropanoid biosynthesis is not clear yet, were also
identified.

Document Number: 176

Environmental pollution is one of the major problems for human health.
Toxic heavy metals are normally present as soil constituents or can also
be spread out in the environment by human activity and agricultural
techniques. Soil contamination by heavy metals as cadmium, highlights
two main aspects: on one side they interfere with the life cycle of
plants and therefore reduce crop yields, and on the other hand, once
adsorbed and accumulated into the plant tissues, they enter the food
chain poisoning animals and humans. Considering this point of view,
understanding the mechanism by which plants handle heavy metal exposure,
in particular cadmium stress, is a primary goal of plant-biotechnology
research or plant breeders whose aim is to create plants that are able
to recover high amounts of heavy metals, which can be used for
phytoremediation, or identify crop varieties that do not accumulate
toxic metal in grains or fruits. In this review we focus on the main
symptoms of cadmium toxicity both on root apparatus and shoots. We
elucidate the mechanisms that plants activate to prevent absorption or
to detoxify toxic metal ions, such as synthesis of phytochelatins,
metallothioneins and enzymes involved in stress response. Finally we
consider new plant-biotechnology applications that can be applied for
phytoremediation.

Document Number: 177

A rigorous safety assessment process exists for GM crops. It includes
evaluation of the introduced protein as well as the crop containing such
protein with the goal of demonstrating the GM crop is "as-safe-as"
non-transgenic crops in the food supply. One of the major issues for GM
crops is the assessment of the expressed protein for allergenic
potential. Currently, no single factor is recognized as an identifier
for protein allergenicity. Therefore, a weight-of-evidence approach,
which takes into account a variety of factors and approaches for an
overall assessment of allergenic potential, is conducted [Codex
Alimentarious Commission, 2003. Alinorm 03/34: Joint FAO/WHO Food
Standard Programme, Codex Alimentarious Commission, Twenty-Fifth
Session, Rome, Italy, 30 June-5 July, 2003. Appendix III, Guideline for
the conduct of food safety assessment of foods derived from
recombinant-DNA plants, and Appendix IV, Annex on the assessment of
possible allergenicity, pp. 47-60]. This assessment is based on what is
known about allergens, including the history of exposure and safety of
the gene(s) source; protein structure (e.g., amino acid sequence
identity to human allergens); stability to pepsin digestion in vitro
[Thomas, K. et al, 2004. A multi-laboratory evaluation of a common in
vitro pepsin digestion assay protocol used in ssessing the safety of
novel proteins. Regul. Toxicol. Pharmacol. 39, 87-98]; an estimate of
exposure of the novel protein(s) to the gastrointestinal tract where
absorption occurs (e.g., protein abundance in the crop, processing
effects); and when appropriate, specific IgE binding studies or skin
prick testing. Additional approaches may be considered (e.g., animal
models; targeted sera screening) as the science evolves; however, such
approaches have not been thoroughly evaluated or validated for
predicting protein allergenicity. (C) 2008 Elsevier Ltd. All Fights
reserved.

Document Number: 178

Proteins introduced into genetically modified (GM) organisms through
genetic engineering must be evaluated for their potential to cause
allergic disease under various national laws and regulations. The Codex
Alimentarius Commission guidance document (2003) calls for testing of
serum IgE binding to the introduced protein if the gene was from an
allergenic source, or the sequence of the transferred Protein has > 35%
identity in any segment of 80 or more amino acids to a known allergen or
shares significant short amino acid identities. The Codex guidance
recognized that the assessment will evolve based on new scientific
knowledge. Arguably, the current criteria are too conservative as
discussed in this paper and they do not provide practical guidance on
serum testing. The goals of this paper are: (I) to summarize evidence
supporting the level of identity that indicates potential risk of
cross-reactivity for those with existing allergies; (2) to provide
example bioinformatics results and discuss their interpretation using
published examples of proteins expressed in transgenic crops; and (3) to
discuss key factors of experimental design and methodology for serum IgE
tests to minimize the rate of false negative and false positive
identification of potential allergens and cross-reactive proteins. (C)
2008 Elsevier Ltd. 

Document Number: 179

Genetic modification of banana has been considered as a path towards
increasing the value of this crop according to health and nutrition in
developing countries. Banana as a crop is one of the most important and
widely consumed fruits as a weaning food by children and as a starchy
staple for all other consumers. As well as providing a low cost and
easily produced source of energy, bananas are also rich in certain
minerals and in vitamins A, C and B6. Growing urbanisation in many
developing countries upgraded the crop importance as a source of
revenue, occasionally providing the main source of income for rural
communities. Genetically modified organism bananas have been advocated
as carrier for vaccines and as a source of carotenoids that can
counteract debilitating vitamin A deficiency. The rather high
vulnerability of banana to pests and diseases triggered biotechnological
applications in an attempt to produce new, more resistant banana
cultivars. However, the potential biosafety of genetically modified
banana and its applications should be taken into account prior to its
extensive usage. The current survey summarises the most important
biotechnological techniques (in vitro culture, DNA fingerprinting,
somatic emrbyogenesis, DNA flow cytometry, etc.) and applications
(micropropagation, in vitro selection, somaclonal variation, protoplast
fusion, haploid production, etc.) in banana and emphasises on genetic
transformation in conjunction with the expressed gene and modified trait
aiming at a further improvement of this crop.

Document Number: 180

Commercial food products from major cities of Coahuila, Mexico were
screened to identify residues of transgenic deoxyribonucleic acid (DNA)
and/or proteins. After performed, an inventory on all products that
contained a soybean-based ingredient in a commercial grocery, store in
the city of Saltillo, Coahuila, Mexico, 245 food products were
identified and grouped in 15 classes according to the soybean ingredient
as well as the manufacturing process used for their elaboration. Similar
sampling was made for The different food classes in the cities of
Monclova, Piedras Negras, and Torreon. A total of 88 samples were
analyzed and DNA was extracted by the hexadecyltrimethyl-ammonium
bromide (CTAB) technique with slight modification to obtain better DNA
quality (1). In addition, segments of the transgenic genes one that
codifies for 5-enolpyruvyshikimate-3-phosphate synthase (epsps), cry 1A,
and the cauliflower mosaic virus (CaMV) promoter were amplified using
polymerase chain reaction (PCR). The transgenic proteins
5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) and insecticidal
crystal protein (Cry 1Ab/Ac) were identified using double antibody
sandwich-enzymatic linked immunoassay analysis (DAS-ELISA). Presence of
transgenic genes and/or proteins was identified in 35.3% of the
commercial products samples.

Document Number: 181

A novel analytical procedure based on the combination of multiplex PCR,
restriction analysis, and CGE-LIF to unambiguosly and simultaneously
confirm the presence of multiple lines of genetically modified corn is
proposed. This methodology is based on the amplification of
event-specific DNA regions by multiplex PCR using 6-FAM-labeled primers.
Subsequently, PCR products are digested by a mixture containing specific
restriction endonucleases. Thus, restriction endonucleases selectively
recognize DNA target sequences contained in the PCR products and cleave
the double-stranded DNA at a given cleavage site. Next, the restriction
digest is analyzed by CGE-LIF corroborating the length of the expected
restriction fragments, confirming (or not) the existence of GMOs. For
accurate size determination of the DNA fragments by CGE-LIF a special
standard DNA mixture was produced in this laboratory for calibration.
The suitability of this mixture for size determination of labeled DNA
fragments is also demonstrated. The usefulness of the proposed
methodology is demonstrated through the simultaneous detection and
confirmatory analysis of samples containing 0.5% of GA21 and MON863
maize plus an endogenous gene of maize as control.

Document Number: 182

Cassava is the staple food of tropical Africa and its production,
averaged over 24 countries, has increased more than threefold from 1980
to 2005, and the population has more than doubled over that time
compared with a 1.5 times increase worldwide. Agriculturally, cassava
performs very well but the roots and leaves contain cyanogenic
glucosides that are dangerous to human health. These cyanogens sometimes
produce acute intoxication leading to death, they exacerbate goitre and
cretinism in iodine-deficient regions, cause konzo and are implicated in
the occurrence of tropical ataxic neuropathy and stunting of children.
Konzo is an irreversible paralysis of the legs with many thousands of
cases, mainly amongst children, in Mozambique, Tanzania, Democratic
Republic of Congo, Cameroon, Central African Republic and probably other
tropical African countries. Attempts to alleviate cassava cyanide
toxicity have included the development of an information network and
distribution in developing countries of picrate kits, which measure
total cyanide in cassava and urinary thiocyanate. A simple wetting
method that reduces total cyanide in cassava flour three- to sixfold has
been successfully field tested and is being introduced in Mozambique.
Transgenic technology shows promise in increasing the rate of loss of
cyanide from roots during processing. World health and agricultural
bodies should pay more attention to emerging health problems associated
with toxicity of cyanogens in cassava. (C) 2008 Society of Chemical
Industry.

Document Number: 183

The International Life Science Institute's Health and Environmental
Sciences Institute's Protein Allergenicity Technical Committee hosted an
international workshop October 23-25, 2007, in Nice, France, to review
and discuss existing and emerging methods and techniques for improving
the current weight-of-evidence approach for evaluating the potential
allergenicity, of novel proteins. The workshop included over 40
international experts from government, industry, and academia. Their
expertise represented a range of disciplines including immunology,
chemistry, molecular biology, bioinformatics, and toxicology. Among
participants, there was consensus that (1) current bioinformatic
approaches are highly conservative; (2) advances in bioinformatics using
structural comparisons of proteins may be helpful as the availability of
structural data increases; (3) proteomics may prove useful for
monitoring the natural variability in a plant's proteome and assessing
the impact of biotechnology transformations on endogenous levels of
allergens, but only when analytical techniques have been standardized
and additional data are available on the natural variation of protein
expression in non-transgenic bred plants: (4) basophil response assays
are promising techniques, but need additional evaluation around
specificity, sensitivity, and reproducibility; (5) additional research
is required to develop and validate an animal model for the purpose of
predicting protein allergenicity. (C) 2008 Elsevier Ltd. All rights
reserved.

Document Number: 184

Plant biotechnology is founded on the principles of cellular totipotency
and genetic transformation, which can be traced back to the Cell Theory
of Matthias Jakob Schleiden and Theodor Schwann, and the discovery of
genetic transformation in bacteria by Frederick Griffith, respectively.
On the 25th anniversary of the genetic transformation of plants, this
review provides a historical account of the evolution of the theoretical
concepts and experimental strategies that led to the production and
commercialization of biotech (transformed or transgenic) plants
expressing many useful genes, and emphasizes the beneficial effects of
plant biotechnology on food security, human health, the environment, and
conservation of biodiversity. In so doing, it celebrates and pays
tribute to the contributions of scores of scientists who laid the
foundation of modern plant biotechnology by their bold and
unconventional thinking and experimentation. It highlights also the many
important lessons to be learnt from the fascinating history of plant
biotechnology, the significance of history in science teaching and
research, and warns against the danger of the growing trends of ignoring
history and historical illiteracy.

Document Number: 185

This paper is the first part of a serial study investigating a
quantification model freed from endogenous reference gene for
genetically modified (GM) content by real-time polymerase chain reaction
(PCR). The serial study involves two parts: (1) quantitative
determination of GM DNA in haplo-species plant samples; (2) quantitative
determination of GM DNA in multi-species plant samples. The paper
describes a methodology to quantify the GM content in a DNA extract
using on one hand real-time PCR to determine the amount of GM targets
present and on the other hand absorbance reading at 260 nm to measure
the total DNA present in sample. The ratio of both values is expressed
as GM percentage. The most prominent dominance of the novel model is
that the direct quantitative relation between the initial amount of
target template in real-time PCR reaction (X(0)) and the content of GM
DNA in tested material (X(SD)) is established. Theoretical analysis
indicates that the developed quantitative model relieved from the
dependence on endogenous reference genes is suitable to quantify the GM
content in haplo-species plant sample, in addition, it has the
applicability in the quantitative detection of GM content in
multi-species GM plant sample. A trail, in which 75 haplo-species GM
plant samples were involved, was conducted to validate the suitability
of the novel quantification model. The bias varied from 0.00 to 24.00%
except a tested sample with lower level of GM content, and the precision
expressed as coefficient of variation (CV) was from 2.81 to 25.00%. The
limit of quantitation (LOQ) of the quantitative assay was as low as
0.1%. Compared with the previous papers and the performance requirements
raised by European Network of GMO Laboratories (ENGL) for analytical
methods of GMO testing, the results demonstrated that the established
quantification model is a suitable alternative to the more traditional
endogenous reference assay in the quantification of GM content in
haplo-species plant sample.

Document Number: 186

Oilseeds are the main source of lipids used in both food and biofuels.
The growing demand for vegetable oil has focused research toward
increasing the amount of this valuable component in oilseed crops.
Globally, soybean (Glycine max) is one of the most important oilseed
crops grown, contributing about 30% of the vegetable oil used for food,
feed, and industrial applications. Breeding efforts in soy have shown
that multiple loci contribute to the final content of oil and protein
stored in seeds. Genetically, the levels of these two storage products
appear to be inversely correlated with an increase in oil coming at the
expense of protein and vice versa. One way to overcome the linkage
between oil and protein is to introduce a transgene that can
specifically modulate one pathway without disrupting the other. We
describe the first, to our knowledge, transgenic soy crop with increased
oil that shows no major impact on protein content or yield. This was
achieved by expressing a codon-optimized version of a diacylglycerol
acyltransferase 2A from the soil fungus Umbelopsis (formerly
Mortierella) ramanniana in soybean seed during development, resulting in
an absolute increase in oil of 1.5% (by weight) in the mature seed.

Document Number: 187

The presence of antibiotic-resistant genes in genetically engineered
crops together with the target gene has generated a number of
environmental and consumer concerns. In order to alleviate public
concerns over the safety of food derived from transgenic crops, marker
gene elimination is desirable. Marker-free transgenic tomato plants were
obtained by using a salicylic-acid-regulated Cre-loxP-mediated
site-specific DNA recombination system in which the selectable marker
neomycin phosphotransferase nptII and cre genes were flanked by two
directly oriented loxP sites. Upon induction by salicylic acid, the cre
gene produced a recombinase that eliminated sequences encoding nptII and
cre genes, sandwiched by two loxP sites from the tomato genome.
Regenerant plants with the Cre-loxP system were obtained by selection on
kanamycin media and polymerase chain reaction (PCR) screening.
Transgenic plants were screened for excision by PCR using nptII, cre,
and PR-1a promoter primers following treatment with salicylic acid. The
footprint of the excision was determined by sequencing the T-DNA borders
after a perfect recombination event. The excision efficiency was 38.7%.
A new plant transformation vector, pBLNSC (Genbank accession number
EU327497), was developed, containing six cloning sites and the
self-excision system. This provided an effective approach to eliminate
the selectable marker gene from transgenic tomato, thus expediting
public acceptance of genetically modified tomato.

Document Number: 188

BACKGROUND: Papaya (Carica papaya L.) production is limited by over 20
viruses, the most damaging of which is papaya ringspot virus (PRSV).
Owing to a lack of suitable PRSV-resistant sources in Carica germplasm,
transgenic resistance using the coat protein (cp) gene of a local PRSV
strain is being developed to manage the disease in Jamaica. For
assurance of food safety, the nutritional and antinutritional
composition of transgenic papayas during ripening was compared with that
of unmodified control samples.
RESULTS: Mature unripe fruits of transgenic and non-transgenic papayas
were repeatedly harvested and stored at room temperature for 1 week
periods, during which random samples were assessed. With the exception
of one transgenic line, no significant differences (P > 0.05) were
observed in selected nutrients and antinutrients between the control and
test samples at three stages of maturity, although a few random
variations were noted.
CONCLUSION: Transformation with viral cp gene and two marker genes did
not produce any major unintended alterations in either the nutritional
or the antinutritional composition of transgenic papayas. These findings
must be compared with other physicochemical and safety assessments to
provide a scientific basis for concluding substantial equivalence with
conventional papayas available on the market in Jamaica. (C) 2008
Society of Chemical Industry.

Document Number: 189

A rapid method was developed for concurrent screening of transgenic
elements in GM canola. This method utilizes a single multiplex PCR
coupled with an oligonucleotide DNA array capable of simultaneously
detecting the 12 approved GM canola lines in Canada. The assay includes
construct-specific elements for identification of approved lines, common
elements (e.g., CaMV 35S promoter, Agrobacterium tumetaciens nos
terminator, or nptII gene) for screening of approved or unapproved
lines, a canola-specific endogenous gene, and endogenous genes from
heterologous crops to serve as additional controls. Oligonucleotide
probes were validated individually for functionality and specificity by
amplification of specific transgene sequences from appropriate GM canola
lines corresponding to each probe sequence, and hybridization of
amplicons to the array. Each target sequence hybridized to its
corresponding oligonucleotide probe and no significant
cross-hybridization was observed. The limit of detection was examined
for the GM lines GT73, T45, and MS8/RF3, and was determined to be 0.1%,
0.1%, and 0.5%, respectively, well within the European food and feed
labeling threshold level of 0.9% for approved GM product. Practically,
the method was demonstrated to be effective for the detection of GM
canola in several types of animal feed, as well as in commercial canola
meal.

Document Number: 190

A diversity of scientific literature supports a role for vitamin D in
decreasing colorectal cancer incidence, but the available evidence
provides only limited support for an inverse association between vitamin
D status and the risk of other types of cancer. We need additional
studies analyzing the dose-response relation between vitamin D status
and cancer risk, the optimal level of 25-hydroxyvitamin D, the length of
time required to observe an effect, and the time period of life when
exposure is most relevant. Studies of vitamin D receptor polymorphisms
have found that not all polymorphisms have the same association with
cancer, and the cancer site could further dictate which polymorphisms
might be most important; this indicates a need for more research on
gene-environment interactions. Several dietary components and the
balance between energy intake and expenditure influence vitamin D
metabolism. These studies show that scientists need to identify
confounders and modifiers of the biological response to vitamin D,
including dietary factors, lifestyle factors such as exercise, and race
or ethnicity. Transgenic and knockout animals are powerful tools for
identifying the molecular targets of bioactive food components.
Scientists should therefore make increased use of these models to
identify molecular targets for vitamin D. Many research gaps relate to
the need to develop predictive, validated, and sensitive biomarkers,
including biomarkers that researchers can use to reliably evaluate
intake or exposure to vitamin D, assess one or more specific biological
effects that are linked to cancer, and effectively predict individual
susceptibility as a function of nutrient-nutrient interactions and
genetics.

Document Number: 191

The effect of heat treatment on the denaturation of Cry1A(b) protein
expressed in transgenic maize was studied over a temperature range of 69
to 77 degrees C. Denaturation of Cry1A(b) protein was measured by the
loss of reactivity with its specific antibodies using a sandwich ELISA.
The process of denaturation was studied by analyzing the values of
inmunoreactive protein after each heat treatment by kinetic analysis.
Denaturation of Cry1A(b) protein was best described assuming a reaction
order of 1.5. D-values calculated were 4338, 2350, 1272, 734, and 601 s
at 69, 71, 73, 75, and 77 degrees C, respectively. Z-value was estimated
to be 9.0 degrees C and the activation energy value was 266.15 kJ/mol.
Thermodynamic parameters for the process of denaturation of Cry1A(b)
protein were also calculated. The high values of the enthalpy of
activation and the positive values of the entropy of activation obtained
for Cry1A(b) protein are typical of a reaction In which the denaturation
of the protein is the rate-determining process that predominates over an
aggregation process during heating.

Document Number: 192

Galactomannan gums are widely used in the food and oil industries, and
there is considerable interest in applying biotechnological approaches
to improve their physical properties. A mannan synthase from guar
(Cyamopsis tetragonoloba) was expressed under the control of a bean
beta-phaseolin promoter in transgenic Medicago truncatula. Although the
expression of exogenous mannan synthase caused a slight decrease in
galactomannan levels in Medicago, the molecular weight and viscosity of
the polymer were significantly increased, although the mannose to
galactose ratio and degree of polydispersity remained unchanged. At the
same time, expression of about 2.8% of the genes was altered
significantly in the seeds of transgenic Medicago lines analysed by
Affymetrix genome chip, with a particularly striking induction of
putative trehalose phosphate synthase genes. Mannan synthase expression
also caused large alterations in the levels of a number of sugars and
sugar alcohols, suggesting that over-expression of a processive
glycosyltransferase perturbs the mechanisms of sugar sensing and/or
homeostasis, possibly involving signalling via trehalose-6-phosphate.

Document Number: 193

The current globally recognized comparative food and feed safety
assessment paradigm for biotechnology-derived crops is a robust and
comprehensive approach for evaluating the safety of both the inserted
gene product and the resulting crop. Incorporating many basic concepts
from food safety, toxicology, nutrition, molecular biology, and plant
breeding, this approach has been used effectively by scientists and
regulatory agencies for 10-15 years. Current and future challenges in
agriculture include the need for improved yields, tolerance to biotic
and abiotic stresses, and improved nutrition. The next generation of
biotechnology-derived crops may utilize regulatory proteins, such as
transcription factors that modulate gene expression and/or endogenous
plant pathways. In this review, we discuss the applicability of the
current safety assessment paradigm to biotechnology-derived crops
developed using modifications involving regulatory proteins. The growing
literature describing the molecular biology underlying plant
domestication and conventional breeding demonstrates the naturally
occurring genetic variation found in plants, including significant
variation in the classes, expression, and activity of regulatory
proteins. Specific examples of plant modifications involving insertion
or altered expression of regulatory proteins are discussed as
illustrative case studies supporting the conclusion that the current
comparative safety assessment process is appropriate for these types of
biotechnology-developed crops. (C) 2008 Elsevier Ltd. All rights
reserved.

Document Number: 194

Specific legislation in the EU requires that foods containing more than
0.9% of genetically modified organisms (GMOs) should be labelled. This
has necessitated the development of methods for detection and
quantification of such materials. Here we present a robust,
quantitative, 9-plex PCR method for event-specific detection of maize
TC1507, MON863, MON810, T25, NK603, GA21, construct specific detection
of BT11, BT176 and detection of the endogenous hmga maize reference
gene. The method is suitable for quantification in the 0-2% range with a
detection limit of approximately 0.1%. PCR is carried out in two stages.
In the first stage, bipartite primers containing a universal 5'-sequence
and a GMO specific 3'-sequence are used. In the second PCR stage only a
universal primer is used. Trypsin digestion between the first and second
PCR stages enhances signal strength and reproducibility. Probes
hybridising to the PCR amplicons are then labelled by primer extension
and detected by fluorescence capillary electrophoresis. Good agreement
was observed in 76 of 80 determinations when 10 food and feed samples
were analysed using the multiplex PCR assay and compared to results from
quantitative real-time 5'-nuclease PCR. The presented method is
therefore suitable for quantification purposes for food and feed
containing the most common maize GMOs.

Document Number: 195

We have developed an efficient system of assessing the ability of a gene
silencing cassette to silence transcripts from recalcitrant or poorly
studied plant species by using a model plant as a host for the gene of
interest. Tobacco plants transgenic for Lachrymatory Factor Synthase
(LFS) enzyme activity from onion were first produced by introducing a
CaMV 35S-onion-lfs gene construct. These plants were then subjected to a
second transformation with an RNAi construct directed against the lfs
gene sequence. LFS enzyme activity assay showed that the transgenic
plants, containing both the lfs gene and the RNAi construct, had
significantly reduced LFS activity. This observation was supported by
Western analysis for the LFS protein and further validated by
quantitative RT-PCR analysis that demonstrated a significant reduction
in the lfs transcript level in the dual transformants. In this work, we
have demonstrated that the RNAi construct is a suitable candidate for
the development of a non-lachrymatory onion. Our model plant RNAi system
has wide-reaching applications for assessment and targeting of plant
secondary pathway genes, from poorly studied or recalcitrant plant
species, that are important in the pharmacological, food and process
industries.

Document Number: 196

Safflower (Carthamus tinctorius L.) is currently being developed as a
platform for the production of novel proteins. Methods for detecting and
quantifying transgenic safflower are needed to ensure seed quality and
to monitor for its adventitious presence. We developed and compared
three methods of assaying for transgenic safflower presence in
conventional seedlots: field bioassays, enzyme-linked immunosorbent
assays (ELISA), and quantitative polymerase chain reaction (Q-PCR).
Limits for reliable quantification for both ELISA and Q-PCR are similar
to 0.1%, although levels at least as low as 0.02% can be detected by
Q-PCR. Levels of quantification for the field bioassay are limited only
by space and resources available. Multiple sampling methods to detect
and quantify transgenic safflower presence at levels lower than 0.1%
were used on field collected samples from a pollen outcrossing
experiment to quantify the adventitious presence of transgenic
safflower. Taking into account the potential utility and relative
advantages or disadvantages of each detection method, it is recommended
that the initial testing for the adventitious presence of transgenic
seed be carried out using an antibody-based test if available and that
Q-PCR-based assays to quantify transgenic proportion be used when it is
necessary to identify specific transgenic constructs or if
antibody-based assays are not readily available.

Document Number: 197

The production of high-value pharmaceutical proteins and peptides in
transgenic plants is an attractive and economically feasible alternative
to conventional mammalian cell, yeast, and bacterial systems. In
contrast to vegetative tissues, rice seeds allow higher accumulation of
recombinant proteins and long-term stable storage. Rice is not only
consumed as a staple food by a majority of the world's population, but
is also used as a model monocot for biotechnological manipulation. Daily
oral consumption of transgenic rice seeds that accumulate high
concentrations of food-derived or synthetic bioactive peptides can be
expected to provide a safe, reliable, and consistent oral delivery
system that would contribute to the promotion of human health care.

Document Number: 198

The objective of this work was to evaluate the proximate composition, as
well as Daidzein (D) and Genistein (G) contents by HPLC, of BRS 243 RR
soybean. Sample preparation for the chromatographic analysis involved
the use of hexane to remove lipids. Isoflavones were extracted with 70%
ethanol containing 0.1% acetic acid. The optimized chromatographic
conditions were: C,,, column, mobile phase methanol:5% acetic acid (1:1
v/v), flow rate 0.5 mL/minute, column temperature 30 degrees C, UV
absorbance at 254 mn and volume injected 40 mu L. The validation
parameters were: linearity of daidzein (y = 11242 x -37433, r = 0.9976)
and genistein (y = 18510 x -6676 1, r = 0.9980); variation coefficients
obtained in intra-day precision assays [CV = 5.3% (D), CV = 6.7% (G) I
and inter-day precision assays [CV = 8.7% (D), CV = 9.7% (G)];
quantification limit 10 mu g.g(-1); detection limit 5 mu g.g(-1) and
recovery 95.7%. Therefore, the optimized method was appropriate for the
determination of daidzein and genistein in soybean. Carbohydrate
(31.4%), protein (35.9%), lipid (20.9%), moisture (6.9%), ash (4.9%),
daidzein (44.1 mu g.g(-1)) and genistein (37.4 mu g.g(-1)) contents
determined in transgenic soybean were similar to those of conventional
soybean determined in other studies.

Document Number: 199

The application of a method based on ligation-dependent probe
amplification to the simultaneous detection of different genetically
modified organisms in food is described. The ligation reaction of
target-specific probes with characteristic lengths and universal primer
binding sites is followed by PCR amplification using fluorescein-labeled
primers. The separation and the detection of DNA fragments are achieved
by capillary electrophoresis via laser-induced fluorescence. The
approach allows the simultaneous detection of several targets
corresponding to different levels of specificity in a one-tube assay.
Synthetic oligonucleotides were designed to detect (1) reference genes
in the genome from maize, soya and rapeseed, (2) the CaMV 35S-promotor
as screening element, (3) the construct-specific 35S-pat junction, and
(4) the event-specific regions of the transgenic maize line MON 810 and
of Roundup Ready soya. Specificity and sensitivity (GMO content 0.1%) of
the approach were shown for all targets. This novel analytical strategy
represents a flexible, modular system for the surveillance of
GMO-derived products that can be readily complemented by further target
sequences of interest.

Document Number: 200

New plant oil crops are desirable as renewable resources for energy, for
food purposes, and as building blocks in chemical synthesis. Fruit oil
of Echinops sphaerocephalus was characterized by a high content of
linoleic acid (over 70% of total fatty acids) and by a high tocopherol
content (530-970 mg/kg oil). The majority was alpha-tocopherol. Echinops
sphaerocephalus plants may be cultivated in agricultural dimensions, and
fruits may be obtained by a combine harvester. Echinops quinoline
alkaloids present a useful by-product of the oil production.

Document Number: 201

Background: Potato is a staple food in the diet of the world's
population and also being used as animal feed. Compared to other crops,
however, potato tubers are relatively poor in the essential amino acid,
methionine. Our aim was to increase the methionine content of tubers by
coexpressing a gene involved in methionine synthesis with a gene
encoding a methionine-rich storage protein in potato plants.
Results: In higher plants, cystathionine gamma-synthase (CgS) is the
first enzyme specific to methionine biosynthesis. We attempted to
increase the methionine content of tubers by expressing the deleted form
of the Arabidopsis CgS (CgS(Delta 90)), which is not regulated by
methionine, in potato plants. To increase the incorporation of free
methionine into a storage protein the CgS(Delta 90) was cotransformed
with the methionine-rich 15-kD beta-zein. Results demonstrated a 2- to
6-fold increase in the free methionine content and in the methionine
content of the zein-containing protein fraction of the transgenic
tubers. In addition, in line with higher methionine content, the amounts
of soluble isoleucine and serine were also increased. However, all of
the lines with high level of CgS(Delta 90) expression were
phenotypically abnormal showing severe growth retardation, changes in
leaf architecture and 40-to 60% reduction in tuber yield. Furthermore,
the colour of the transgenic tubers was altered due to the reduced
amounts of anthocyanin pigments. The mRNA levels of phenylalanine
ammonia-yase (PAL), the enzyme catalysing the first step of anthocyanin
synthesis, were decreased.
Conclusion: Ectopic expression of CgS(Delta 90) increases the methionine
content of tubers, however, results in phenotypic aberrations in potato.
Co-expression of the 15-kD P-zein with CgS(Delta 90) results in
elevation of protein-bound methionine content of tubers, but can not
overcome the phenotypical changes caused by CgS(Delta 90) and can not
significantly improve the nutritional value of tubers. The level of PAL
mRNA and consequently the amount of anthocyanin pigments are reduced in
the CgS(Delta 90) transgenic tubers suggesting that methionine synthesis
and production of anthocyanins is linked.

Document Number: 202

Genetic modification of crop plants to introduce desirable traits such
as nutritional enhancement, disease and pest resistance, and enhanced
crop productivity is increasingly seen as a promising technology for
sustainable agriculture and boosting food production in the world.
Independently, cultural practices that utilize alternative agriculture
strategies including organic cultivation subscribe to sustainable
agriculture by limiting chemical usage and reduced tillage. How the two
together affect fruit metabolism or plant growth in the field or whether
they are compatible has not yet been tested. Fruit-specific yeast
S-adenosylmethionine decarboxylase (ySAMdc) line 579HO, and a control
line 556AZ were grown in leguminous hairy vetch (Vicia villosa Roth)
(HV) mulch and conventional black polyethylene (BP) mulch, and their
fruit analysed. Significant genotypexmulch-dependent interactions on
fruit phenotype were exemplified by differential profiles of 20 fruit
metabolites such as amino acids, sugars, and organic acids. Expression
patterns of the ySAMdc transgene, and tomato SAMdc, E8, PEPC, and ICDHc
genes were compared between the two lines as a function of growth on
either BP or HV mulch. HV mulch significantly stimulated the
accumulation of asparagine, glutamate, glutamine, choline, and citrate
concomitant with a decrease in glucose in the 556AZ fruits during
ripening as compared to BP. It enables a metabolic system in tomato
somewhat akin to the one in higher polyamine-accumulating transgenic
fruit that have higher phytonutrient content. Finally, synergism was
found between HV mulch and transgenic tomato in up-regulating N:C
indicator genes PEPC and ICDHc in the fruit.

Document Number: 203

Agrobacterium-mediated transformation for poinsettia (Euphorbia
pulcherrima Willd. Ex Klotzsch) is reported here for the first time.
Internode stem explants of poinsettia cv. Millenium were transformed by
Agrobacterium tumefaciens, strain LBA 4404, harbouring virus-derived
hairpin (hp) RNA gene constructs to induce RNA silencing-mediated
resistance to Poinsettia mosaic virus (PnMV). Prior to transformation,
an efficient somatic embryogenesis system was developed for poinsettia
cv. Millenium in which about 75% of the explants produced somatic
embryos. In 5 experiments utilizing 868 explants, 18 independent
transgenic lines were generated. An average transformation frequency of
2.1% (range 1.2-3.5%) was revealed. Stable integration of transgenes
into the poinsettia nuclear genome was confirmed by PCR and Southern
blot analysis. Both single- and multiple-copy transgene integration into
the poinsettia genome were found among transformants. Transgenic
poinsettia plants showing resistance to mechanical inoculation of PnMV
were detected by double antibody sandwich enzyme-linked immunosorbent
assay (DAS-ELISA). Northern blot analysis of low molecular weight RNA
revealed that transgene-derived small interfering (si) RNA molecules
were detected among the poinsettia transformants prior to inoculation.
The Agrobacterium-mediated transformation methodology developed in the
current study should facilitate improvement of this ornamental plant
with enhanced disease resistance, quality improvement and desirable
colour alteration. Because poinsettia is a non-food, non-feed plant and
is not propagated through sexual reproduction, this is likely to be more
acceptable even in areas where genetically modified crops are currently
not cultivated.

Document Number: 204

beta-Amylase hydrolyzes the alpha-1,4-glycosidic linkages of starch
resulting in the release of maltose. This reaction is of industrial
importance for maltose production and for the preparation process of
fermented foods and alcoholic beverages. A demand for an acceleration of
the rate of enzymatic cleavage of the starch macro-molecule is a
prerequisite for large-scale and highly efficient production. Increasing
the temperature up to the optimum of approximately 60 degrees C can
significantly speed up the reaction. However, at higher temperatures,
the effect on protein denaturation becomes dominant, and the conversion
rate decreases. The primary objective of this study was to generate
transgenic plants of the "Kennebec" potato variety for production of
thermostable beta-amylase using Agrobacterium-mediated transformation.
Four chimeric genes encoding the beta-amylase with or without signal
peptide sequences for targeting expression in cytoplasm, amyloplasts, or
vacuoles were constructed and driven by high tuber expression promoter
from Sucrose synthetase gene Sus4. Forty-two transgenic lines were
selected for this study. Transgenic lines with various beta-amylase
constructs were verified for the existence and expression of the
transgenes by PCR approaches. The expression level of the introduced
beta-amylase protein was estimated by immunoblot analyses using
polyclonal antibodies. Recombinant beta-amylase was successfully
expressed in Escherichia coli B21 (DE3), and temperature ranges of these
inducible recombinant proteins were found to be between 40 and 90
degrees C. This enzymatic complex produced in the in vitro cultured
microtubers and field-grown tubers from transgenic potatoes were proved
to be stable and active at 60 degrees C. The relative activities of
beta-amylase in tubers of field-grown potatoes were compared, and the
maximum increase was found with transgenic line #6A of the pSUS4-AMY
construct which has an 11 fold greater increase than the untransformed
"Kennebec". Variations of the chemical compositions were found in the
selected transgenic lines. Results of this study suggest the feasibility
of utilizing thermostable beta d-amylase in transgenic potatoes for the
starch-processing industries. (C) 2008 Elsevier Ireland Ltd. All rights
reserved.

Document Number: 205

Oxalate, a common constituent in many plants, is known to play important
functional roles in plants. However, excess levels of oxalate in edible
parts of plants adversely affect their quality as food. Understanding
the regulatory mechanism in plants, particularly in food crops, is of
both scientific and practical significance. While a number of studies
have shown that nitrate can efficiently induce oxalate accumulation in
plants, how it elicits such an effect is not well understood. This study
aimed to gain a further insight into the mechanism underlying the
nitrate-induced oxalate accumulation. Nitrate-N efficiently caused
oxalate accumulation in rice leaves, depending on the nitrate
concentrations and treatment time. In contrast, same nitrogen molar
levels of the other N forms such as nitrite, ammonium, glutamate and
urea either had no effect on the accumulation or even reduced the
oxalate level. When glutamate, glutamine, asparate and asparagine were
added into the nutrient solution that already contained saturating
concentration of nitrate, both oxalate levels and NR activity were
correspondingly decreased. In all of these modes of treatment, the
change in NR activity was positively paralleled to that in oxalate
levels. For a further confirmation, we generated the transgenic rice
plants with a NR interference gene introduced. The result further
demonstrated that in the transgenic plants, unlike in wild-type plants,
oxalate was no longer able to accumulate in response to the nitrate
treatment even though the endogenous nitrate levels were substantially
elevated. Taken together, our results suggest that the nitrate-induced
oxalate accumulation in rice leaves is dependent on the NR-catalyzed
nitrate reduction, rather than on nitrate itself or nitrite reduction or
its downstream metabolites.

Document Number: 206

Plants produce a range of biopolymers for purposes such as maintenance
of structural integrity, carbon storage, and defense against pathogens
and desiccation. Several of these natural polymers are used by humans as
food and materials, and increasingly as an energy carrier. In this
review, we focus on plant biopolymers that are used as materials in bulk
applications, such as plastics and elastomers, in the context of
depleting resources and climate change, and consider technical and
scientific bottlenecks in the production of novel or improved materials
in transgenic or alternative crop plants. The biopolymers discussed are
natural rubber and several polymers that are not naturally produced in
plants, such as polyhydroxyalkanoates, fibrous proteins and poly-amino
acids. In addition, monomers or precursors for the chemical synthesis of
biopolymers, such as 4-hydroxybenzoate, itaconic acid, fructose and
sorbitol, are discussed briefly.

Document Number: 207

One component of the safety assessment of agricultural products produced
through biotechnology is evaluation of the safety of newly expressed
proteins. The ILSI International Food Biotechnology Committee has
developed a scientifically based two-tiered, weight-of-evidence strategy
to assess the safety of novel proteins used in the context of
agricultural biotechnology. Recommendations draw upon knowledge of the
biological and chemical characteristics of proteins and testing methods
for evaluating potential intrinsic hazards of chemicals. Tier I
(potential hazard identification) includes an assessment of the
biological function or mode of action and intended application of the
protein, history of safe use, comparison of the amino acid sequence of
the protein to other proteins, as well as the biochemical and
physico-chemical properties of the proteins. Studies outlined in Tier II
(hazard characterization) are conducted when the results from Tier I are
not sufficient to allow a determination of safety (reasonable certainty
of no harm) on a case-by-case basis. These studies may include acute and
repeated dose toxicology studies and hypothesis-based testing. The
application of these guidelines is presented using examples of
transgenic proteins applied for agricultural input and output traits in
genetically modified crops along with recommendations for future
research considerations related to protein safety assessment. (C) 2008
ILSI. Published by Elsevier Ltd. 

Document Number: 208

Genetically modified transgenic foods (TF) have generated considerable
debate in developed countries. Therefore, a survey of 400 people was
conducted to assess the importance of this variable in consumer
decisions regarding the purchase of liquid milk and tomato sauce in
Temuco (IX(th) Region, Chile). This survey also evaluated the existence
of different market segments. Using a conjoint analysis, we established
that the presence of genetic modification (GM) in food was a more
important factor than either the brand or the price in both types of
food. Using a cluster analysis, five consumer groups were distinguished,
with two large groups representing 46.8% and 18.8% of the total number
of people surveyed. These two groups felt the existence of GMs to be
important, but only the largest group preferred the alternatives with no
GMs. For the third group (16.8%), brand was the most important
attribute, but a preference for non-TF was also shown. The fourth group
(10.5%) placed great importance on price and preferred non-TE The
smallest group (7.3%) placed a slightly higher value on the price of
milk and on the brand of tomato sauce, showing a preference for TF The
five groups all rejected the store-brand product and most consumers
reacted positively to lower prices. Therefore, with the results of this
investigation, it can be concluded that the absence of GMs in milk and
tomato sauce is a desirable condition for a large proportion of
consumers in Temuco (74%), but there are groups of consumers (26%) that
accept genetic manipulation of these foods.

Document Number: 209

Qualitative and quantitative polymerase chain reaction-based methods to
detect genetically modified soy (RoundupReady (TM) soy) and maize (Bt176
Maximizer (TM) maize, MON810 YieldGard (TM) corn, T25 LibertyLink (TM)
maize) were applied to processed foods sold commercially in Brazil. From
2000 to 2005, 100 food products containing maize and 100 food products
containing soy were analysed every single year. The presence of
genetically modified soy increased steadily from 13% in 2000 to 78% in
2005. The number of food products containing genetically modified soy in
a proportion above 1.0% on the ingredient level, the threshold for
labelling according to Brazilian legislation, increased from 11% in 2000
to 36%,, in 2005. No clear trend was found within maize containing food
products. Eight to eleven percent were shown to consist of material
derived from genetically modified maize and 4-6% were found to contain
more than 1% of genetically modified maize. (c) 2007 Elsevier Ltd. All
rights reserved.

Document Number: 210

Transgenesis provides a method of expressing novel proteins in milk to
increase the functional benefits of milk consumption. Transgenic goats
expressing human lysozyme (hLZ) at 67% of the concentration in human
breast milk were produced, thereby enhancing the antimicrobial
properties of goats' milk. The objective of this study was to
investigate the impact of pasteurized milk containing hLZ on growth, the
intestinal epithelium, and an enteropathogenic Escherichia coli (EPEC)
infection in young weaned pigs. Pigs were placed into 4 groups and fed a
diet of solid food and either control (nontransgenic) goats' milk or
milk from hLZ-transgenic goats, Growth was assessed by weight gain.
Nonchallenged pigs were necropsied after 6 wk, whereas the remaining
pigs were necropsied at 7 wk following bacterial challenge. We
determined the numbers of total coliforms and E. coli and examined small
intestinal histology for ail pigs. Complete blood counts were also
determined pre- and postchallenge. Challenged pigs receiving hLZ milk
had fewer total coliforms (P = 0.029) and E. coli(P = 0.030) in the
ileum than controls. hLZ-fed pigs also had a greater duodenal villi
width (P = 0.029) than controls. Additionally, nonchallenged hLZ-fed
pigs had fewer intraepithelial lymphocytes per micron of villi height (P
= 0.020) than nonchallenged controls. These results indicate that the
consumption of pasteurized hLZ goats' milk has the potential to improve
gastrointestinal health and is protective against an EPEC in young
weaned pigs. These same benefits may occur in young children if they
were to consume milk from hLZ-transgenic goats.

Document Number: 211

The Interregional Research Project Number 4 (IR-4) Specialty Food Crops
Program is a publicly-funded program initiated in 1963 to develop and
submit regulatory data to support registration of pest control products
for specialty crops. In the early to mid 1990s, nearly 45% of the IR-4
residue projects supported new herbicide registrations for fruits and
vegetables with the other 55% devoted to fungicides, insecticides, and
nematacides. In 2005, the number of residue projects conducted by IR-4
to support herbicide fruit and vegetable registrations was less than
30%. The three main factors that have contributed to this decline are:
fewer herbicides available for registration; product liability concerns;
and an increased focus on new, safer, and Reduced Risk Pesticides for
insect and disease control. It has been a number of years since a new
herbicide has been developed for a major crop that could be extended to
specialty food crops. Many of the current IR-4 herbicide projects are
with products that have been on the market for 20 or more years. Product
liability is a concern because of the high value of many specialty crops
relative to the potential market opportunity. In many cases, the
registrant requires product performance data before IR-4 can proceed
with a residue project. With limited funds for developing these data,
many new projects never proceed to the regulatory stage. Although
registrants can seek indemnification for some of these uses, it is a
complicated often state-specific process. IR-4 has been successful in a
number of areas, including the registration of a large numbers of uses
through reduced data extrapolations for products such as glyphosate and
carfentrazone-ethyl. Additionally, IR-4 submitted the first successful
petition establishing an exemption of tolerance for a conventional
herbicide (imazamox). Future IR-4 initiatives include collaboration with
industry, growers, and academia to develop new herbicide technologies
such as plant breeding or transgenic crops and generation of appropriate
data to extend those products to specialty food crops. IR-4 will also
assist in registering products that can be used on crops that have been
selected (or developed through biotechnological approaches) to be
tolerant to existing herbicides. Registrants should strongly consider
developing herbicides for specialty food crops, with IR-4's assistance,
as a means to expand markets and also as a means to extend data
protection of their products, as allowed under the Food Quality
Protection Act. Nomenclature: Carfentrazone-ethyl; glyphosate; imazamox.

Document Number: 212

Potato (Solanum tuberosum L.), one of the most important food crops, is
susceptible to a number of devastating fungal pathogens in addition to
bacterial and other pathogens. Producing disease-resistant cultivars has
been an effective and useful strategy to combat the attack of pathogens.
Potato was transformed with Agrobacterium tumefaciens strain EHA101
harboring chitinase, (ChiC) isolated from Streptomyces griseus strain
HUT 6037 and bialaphos resistance ( bar) genes in a binary plasmid
vector, pEKH1. Polymerase chain reaction (PCR) analysis revealed that
the ChiC and bar genes are integrated into the genome of transgenic
plants. Different insertion sites of the transgenes ( one to six sites
for ChiC and three to seven for bar) were indicated by Southern blot
analysis of genomic DNA from the transgenic plants. Expression of the
ChiC gene at the messenger RNA ( mRNA) level was confirmed by Northern
blot analysis and that of the bar gene by herbicide resistance assay.
The results obviously confirmed that the ChiC and bar genes are
successfully integrated and expressed into the genome, resulting in the
production of bialaphos-resistant transgenic plants. Disease-resistance
assay of the in vitro and greenhouse-grown transgenic plants
demonstrated enhanced resistance against the fungal pathogen Alternaria
solani ( causal agent of early blight).

Document Number: 213

Salinity stress is a major limiting factor in cereal productivity. Many
studies report improvements in salt tolerance using model plants, such
as Arabidopsis thaliana or standard varieties of rice, e. g., the
japonica rice cultivar Nipponbare. However, there are few reports on the
enhancement of salt tolerance in local rice cultivars. In this work, we
used the indica rice ( Oryza sativa) cultivar BR5, which is a local
cultivar in Bangladesh. To improve salt tolerance in BR5, we introduced
the Escherichia coli catalase gene, katE. We integrated the katE gene
into BR5 plants using an Agrobacterium tumefaciens-mediated method. The
introduced katE gene was actively expressed in the transgenic BR5 rice
plants, and catalase activity in T-1 and T-2 transgenic rice was
approximately 150% higher than in nontransgenic plants. Under NaCl
stress conditions, the transgenic rice plants exhibited high tolerance
compared with nontransgenic rice plants. T2 transgenic plants survived
in a 200 mM NaCl solution for 2 weeks, whereas nontransgenic plants were
scorched after 4 days soaking in the same NaCl solution. Our results
indicate that the katE gene can confer salt tolerance to BR5 rice
plants. Enhancement of salt tolerance in a local rice cultivar, such as
BR5, will provide a powerful and useful tool for overcoming food
shortage problems.

Document Number: 214

Polyphenol oxidases (PPOs), which catalyze the oxidation of phenolics to
quinones, have been reported to confer resistance to Pseudomonas
syringae and are thought to be involved in insect resistance. To assess
the impact of PPO expression on resistance to the common cutworm
(Spodoptera litura (F.)) (Lepidoptera: Noctuidae), we used transgenic
tomato (Lycopersicon esculentum Mill.) plants constitutively expressing
sense- and antisense-oriented potato PPO genes. Transgenic plants
expressing a sense PPO construct (overexpressing PPO [OP] plants)
exhibited 2.0-5.7-fold higher PPO activity levels, whereas antisense PPO
transgenic plants (suppressed PPO [SP] plants) exhibited 1.5-7.3-fold
lower PPO activity levels than nontransformed controls. The
PPO-overexpressing transgenic plants clearly showed an increase in
resistance; simple growth rates of common cutworms on OP plants were up
to 2.5 and 3.3 times lower than on controls and SP leaves, respectively,
and larvae consumed less foliage. In addition, increased PPO activity
led to higher larval mortality. The efficiency of conversion of ingested
food and efficiency of conversion of digested food of third instars were
found to be significantly different among tomato genotypes with
differing PPO activity levels, suggesting that PPO activity rendered
foliage less nutritious. Moreover, when leaflets at nodes 4 and 8 of SP,
NT and OP plants were detached and fed to common cutworm larvae, their
PPO activity levels were induced 1.6-2.2-fold. These results suggest a
critical role for PPO-mediated phenolic oxidation in resistance to this
insect. Manipulation of PPO activity could provide resistance
simultaneously to both disease and insect pests, and therefore might be
used as a component of effective integrated pest management. (C) 2008
Elsevier Ireland Ltd. 

Document Number: 215

Micronutrient malnutrition affects more than half of the world
population, particularly in developing countries. Concerted
international and national fortification and supplementation efforts to
curb the scourge of micronutrient malnutrition are showing a positive
impact, alas without reaching the goals set by international
organizations. Biofortification, the delivery of micronutrients via
micronutrient-dense crops, offers a cost-effective and sustainable
approach, complementing these efforts by reaching rural populations.
Bioavailable micronutrients in the edible parts of staple crops at
concentrations high enough to impact on human health can be obtained
through breeding, provided that sufficient genetic variation for a given
trait exists, or through transgenic approaches. Research and breeding
programs are underway to enrich the major food staples in developing
countries with the most important micronutrients: iron, provitamin A,
zinc and folate.

Document Number: 216

Seed tissues (endosperm, embryo, and pericarp) are often separated into
tissue-enriched fractions by wet- or dry-milling methods for use in
food, feed, and industrial products. Seed tissue markers that are
sensitive and readily quantifiable would be useful to optimize
fractionation processes. To meet this need for tissue markers, we set
out to produce and characterize different transgenic maize lines, each
containing green fluorescent protein (GFP) in either endosperm or
embryo. We examined mRNA transcripts using expressed sequence tag (EST)
profiles of several major seed proteins and selected several with strong
seed tissue preferences. Stably transformed maize lines were produced,
and visual observation of fluorescence confirmed the presence of GFP in
the desired tissues. To establish the utility of this grain for
evaluating the effectiveness or separation efficiencies of fractionation
processes, transgenic kernels were hand-dissected into pericarp,
endosperm, and embryo fractions and the GFP concentration in each
fraction was determined. The GFP distribution between fractions of each
transgenic event was calculated from GFP concentration and mass balance,
which enabled the determination of GFP yield based on the
hand-dissection fractionation data and the amount of tissue
contamination in each fraction. Our transgenic lines exhibited strong
tissue preference for either embryo or endosperm. These lines should be
useful for assessing separation efficiencies in maize fractionation
processes.

Document Number: 217

Petroleum dependency is a challenge that can potentially be partly
offset by agricultural production of biofuels, while decreasing net,
nonrenewable carbon dioxide output. Plants have not been domesticated
for modern biofuel production, and the quickest, most efficient, and
often, the only way to convert plants to biofuel feedstocks is
biotechnologically. First generation biofuel feedstock sources:
sugarcane and cereal grains to produce bioethanol and biobutanol and
oilseeds to produce biodiesel compete directly with needs for world food
security. The heavy use of oilseed rape releases quantities of methyl
bromide to the atmosphere, which can be prevented by gene suppression.
Second generation bioethanolic/biobutanolic biofuels will come from
cultivated lignocellulosic crops or straw wastes. These presently
require heat and acid to remove lignin, which could be partially
replaced by transgenically reducing or modifying lignin content and
upregulating cellulose biosynthesis. Non-precipitable silicon emissions
from burning could be reduced by transgenically modulating silicon
content. The shrubby Jatropha and castor beans should have highly toxic
protein components transgenically removed from their meal, cancer
potentiating diterpenes removed from the oils, and allergens from the
pollen, before extensive cultivation. Algae and cyanobacteria for third
generation biodiesel need transgenic manipulation to deal with "weeds",
light penetration, photoinhibition, carbon assimilation, etc. The
possibilities of producing fourth generation biohydrogen and
bioelectricity using photosynthetic mechanisms are being explored. There
seem to be no health or environmental impact study requirements when the
undomesticated biofuel crops are grown, yet there are illogically
stringent requirements should they transgenically be rendered less toxic
and more efficient as biofuel crops. (c) 2007 Elsevier Ireland Ltd. All
rights reserved.

Document Number: 218

The present study considers the debate generated in developed countries
by genetically modified foods, the importance of this variable to
consumers in Temuco (Araucania Region, Chile) when purchasing tomato
sauce and different market segments were studied through a personal
survey administered to 400 people. Using conjoint analysis, it was
determined that the presence of genetic modification in food was
generally more important than the brand and purchase price. Using
cluster analysis, three segments were distinguished, with the most
numerous (49.3%) placing the greatest importance on the presence of
genetic modification (GM) in food and rejecting the transgenic product.
The second group (39.4%) gave the greatest importance to the brand and
preferred tomato sauce with genetically modified ingredients. The
smallest segment (11.3%) placed the greatest value on price and
preferred transgenic tomato sauce. The three segments prefer the
national brand, reject the store brand and react positively to lower
prices. The segment sensitive to the presence of GM in food comprised
mainly those younger than 35 years of age, single and with no children.
The absence of GM in food of vegetable origin is desirable for young
consumers in the Araucania Region, but a significant proportion accepts
genetic modification in food (50.7%).

Document Number: 219

Peanut allergy is one of the most life-threatening food allergies and
one of the serious challenges facing the peanut and food industries.
Current proposed solutions focus primarily on ways to alter the immune
system of patients allergic to peanut. However, with the advent of
genetic engineering novel strategies can be proposed to solve the
problem of peanut allergy from the source. The objectives of this study
were to eliminate the immunodominant Ara h 2 protein from transgenic
peanut using RNA interference (RNAi), and to evaluate the allergenicity
of resulting transgenic peanut seeds. A 265-bp-long PCR product was
generated from the coding region of Ara h 2 genomic DNA, and cloned as
inverted repeats in pHANNIBAL, an RNAi-inducing plant transformation
vector. The Ara h 2-specific RNAi transformation cassette was subcloned
into a binary pART27 vector to construct plasmid pDK28. Transgenic
peanuts were produced by infecting peanut hypocotyl explants with
Agrobacterium tumefaciens EHA 105 harbouring the pDK28 construct. A
total of 59 kanamycin-resistant peanut plants were regenerated with
phenotype and growth rates comparable to wild type. PCR and Southern
analyses revealed that 44% of plants stably integrated the transgene.
Sandwich ELISA performed using Ara h 2-mAbs revealed a significant (P <
0.05) reduction in Ara h 2 content in several transgenic seeds. Western
immunobloting performed with Ara h 2-mAb corroborated the results
obtained with ELISA and showed absence of the Ara h 2 protein from crude
extracts of several transgenic seeds of the T-0 plants. The
allergenicity of transgenic peanut seeds expressed as IgE binding
capacity was evaluated by ELISA using sera of patients allergic to
peanut. The data showed a significant decrease in the IgE binding
capacity of selected transgenic seeds compared to wild type, hence,
demonstrating the feasibility of alleviating peanut allergy using the
RNAi technology.

Document Number: 220

This study shows that a diet including insect-resistant Bt176 maize, fed
to 53 ewes and their progeny for 3 years, did not have adverse effects
on their health or performance and that no horizontal gene transfer to
ruminal microorganisms or animal tissues was detected. No differences
were observed regarding performance, reproductive traits, haematological
parameters, antioxidant defences, lymphocyte proliferative capacity,
phagocytosis and intracellular killing of macrophages, and ruminal
microbial population characteristics between control and genetically
modified (GM) maize-fed animals. Immune response to Salmonella abortus
ovis vaccination was more efficient in GM maize fed sheep. No
modifications of histological features of tissues were found; however,
cytochemical analyses of ruminal epithelium by Ki67 staining provided
evidence of proliferative activation of basal cells in all GM maize-fed
ewes. Preliminary electron microscopy analyses of the liver and pancreas
revealed smaller cell nuclei containing increased amounts of
heterochromatin and perichromatin granules in GM maize-fed lambs. Meat
protein content and water loss by cooking were slightly affected by the
dietary treatment. No transgenic DNA was detected in tissues, blood, and
ruminal fluid or ruminal bacteria. Longitudinal studies should be
included in evaluation of food safety whenever possible and sheep may be
a useful animal model for toxicological assessment. (C) 2007 Elsevier
B.V. 

Document Number: 221

Ms8xRf3 is a genetically modified rapeseed hybrid which is widely
cultivated in Canada and exported to some other countries for production
of foodstuffs or fodder. In this study, the genomic sequences flanking
the right borders of the integrated transgenic sequences in the Ms8xRf3
genome were characterized and showed high similarities with the
bacterial artificial chromosome clone of Chinese cabbage. Event-specific
qualitative polymerase chain reaction (PCR) methods were established
with the primers and probes targeting the junction regions to produce a
123 base pair (bp) product for the Ms8 event and 92 bp for the Rf3
event. The absolute detection limit of qualitative PCR was 2.5 initial
template copies for the Ms8 event and 50 copies for the Rf3 event.
Quantitiative detection methods were established, with the absolute
quantification limit being approximately 25 initial template copies.

Document Number: 222

This paper reviews the history of the federal regulatory oversight of
plant agricultural biotechnology in the USA, focusing on the scientific
and political forces moulding the continually evolving regulatory
structure in place today. Unlike most other jurisdictions, the USA
decided to adapt pre-existing legislation to encompass products of
biotechnology. In so doing, it established an overarching committee
(Office of Science and Technology Policy) to study and distribute
various regulatory responsibilities amongst relevant agencies: the Food
and Drug Administration, Environmental Protection Agency and US
Department of Agriculture. This paper reviews the history and procedures
of each agency in the execution of its regulatory duties and
investigates the advantages and disadvantages of the US regulatory
strategy.

Document Number: 223

gamma-Linolenic acid (GLA; C18:3 Delta 6,9,12) is a nutritionally
important fatty acid (FA) playing a vital role in biological structures
and cellular functions, which is not produced in oil seed crops. Many
oil seed plants, however, produce significant quantities of linoleic
acid, a FA that could be converted into GLA by the enzyme Delta 6
desaturase, if it is present. As a first step to produce GLA in oil seed
crops, we isolated a cDNA encoding the Delta 6-FA desaturase from
filamentous fungus Mucor circinelloides M29. Expression of this gene in
transgenic tobacco resulted in the accumulation of GLA to the levels of
23.1% of the total FA. The results suggested that it is feasible to
introduce the M. circinelloides Delta 6 desaturase gene into
conventional oil crop to produce a large amount of GLA for functional
foods and pharmaceutical products.

Document Number: 224

Zinc deficiency is a well-documented problem in food crops, causing
decreased crop yields and nutritional quality. Generally, the regions in
the world with Zn-deficient soils are also characterized by widespread
Zn deficiency in humans. Recent estimates indicate that nearly half of
world population suffers from Zn deficiency. Cereal crops play an
important role in satisfying daily calorie intake in developing world,
but they are inherently very low in Zn concentrations in grain,
particularly when grown on Zn-deficient soils. The reliance on
cereal-based diets may induce Zn deficiency-related health problems in
humans, such as impairments in physical development, immune system and
brain function. Among the strategies being discussed as major solution
to Zn deficiency, plant breeding strategy (e.g., genetic
biofortification) appears to be a most sustainable and cost-effective
approach useful in improving Zn concentrations in grain. The breeding
approach is, however, a long-term process requiring a substantial effort
and resources. A successful breeding program for biofortifying food
crops with Zn is very much dependent on the size of plant-available Zn
pools in soil. In most parts of the cereal-growing areas, soils have,
however, a variety of chemical and physical problems that significantly
reduce availability of Zn to plant roots. Hence, the genetic capacity of
the newly developed (biofortified) cultivars to absorb sufficient amount
of Zn from soil and accumulate it in the grain may not be expressed to
the full extent. It is, therefore, essential to have a short-term
approach to improve Zn concentration in cereal grains. Application of Zn
fertilizers or Zn-enriched NPK fertilizers (e.g., agronomic
biofortification) offers a rapid solution to the problem, and represents
useful complementary approach to on-going breeding programs. There is
increasing evidence showing that foliar or combined soil+foliar
application of Zn fertilizers under field conditions are highly
effective and very practical way to maximize uptake and accumulation of
Zn in whole wheat grain, raising concentration up to 60 mg Zn kg(-1).
Zinc-enriched grains are also of great importance for crop productivity
resulting in better seedling vigor, denser stands and higher stress
tolerance on potentially Zn-deficient soils. Agronomic biofortification
strategy appears to be essential in keeping sufficient amount of
available Zn in soil solution and maintaining adequate Zn transport to
the seeds during reproductive growth stage. Finally, agronomic
biofortification is required for optimizing and ensuring the success of
genetic biofortification of cereal grains with Zn. In case of greater
bioavailability of the grain Zn derived from foliar applications than
from soil, agronomic biofortification would be a very attractive and
useful strategy in solving Zn deficiency-related health problems
globally and effectively.

Document Number: 225

Sugar beet (Beta vulgaris L.) is an important industrial crop, being one
of only two plant sources from which sucrose (i.e., sugar) can be
economically produced. Despite its relatively short period of
cultivation (ca. 200 years), its yield and quality parameters have been
significantly improved by conventional breeding methods. However, during
the last two decades or so, advanced in vitro culture and genetic
transformation technologies have been incorporated with classical
breeding programs, the main aim being the production of herbicide-and
salt-tolerant, disease- and pest-resistant cultivars. Among the many
applications of in vitro culture techniques, sugar beet has benefited
the most from haploid plant production, protoplast culture, and
somaclonal variation and in vitro cell selection. Several genetic
transformation technologies have been developed, such as
Agrobacterium-meditated, PEG-mediated, particle bombardment,
electroporation, sonication and somatic hybridization, the first two
being the most successful. Development of herbicide- and salt-tolerant,
virus-, pest/nematode-, fungus/Cercospora- and insect-resistant sugar
beet has been demonstrated. However, only herbicide-tolerant varieties
have been approved for commercialization but not yet available in the
marketplace; rhizomania-resistant varieties are being evaluated in field
trials. Transgenic plants that convert sucrose into fructan, a polymer
of fructose, were also developed. Initial attempts to increase sucrose
yields produced promising results, but it still requires additional
work. Despite marked progress in improving regeneration and
transformation of sugar beet, genotype dependence and low regeneration
and transformation frequencies are still serious restrictions for
routine application of in vitro culture and, more importantly,
transformation technologies. Selected food safety and environmental
impact, as well as regulatory and public acceptance issues relating to
transgenic sugar beet are also discussed.

Document Number: 226

The Rab GTPases are important components of endocytic network in plant
cells. Endocytosis participates in the cell's reaction to extracellular
stimuli by desensitizing, down-regulating or recycling receptors and
membrane proteins. Rab7 is a small GTP-binding protein involved in
intracellular vesicle trafficking from late endosome to the vacuole. We
have isolated Rab7 cDNA from Pennisetum glaucum, a relatively
drought-stress tolerant food grain crop grown commonly in India, during
cDNA-subtractive hybridization of dehydration-stress treated plants. The
PgRab7 ORF, encoding 207 aminoacids, was over-expressed in E. coli. The
recombinant PgRab7 protein showed GTP-binding and GTPase activity.
Transcript expression of PgRab7 gene was differentially up-regulated by
different environmental stimuli such as cold, dehydration and NaCl and
also by a plant hormone IAA. Overexpression of PgRab7 gene enhanced
tolerance to NaCl and mannitol in transgenic tobacco. Transgenic plants
also had increased alkaline phosphatase (ALP) activity. These results
show that PgRab7 is a potential candidate gene for developing both
salinity and dehydration tolerance in planta.

Document Number: 227

This paper discusses the ethical and regulatory issues raised by
"intragenics" - organisms that have been genetically modified using gene
technologies, but that do not contain DNA from another species.
Considering the rapid development of knowledge about gene regulation and
genomics, we anticipate rapid advances in intragenic methods. Of
regulatory systems developed to govern genetically modified organisms
(GMOs) in North America, Europe, Australia, and New Zealand, the
Australian system stands out in explicitly excluding intragenics from
regulation. European systems are also under pressure to exclude
intragenics from regulation. We evaluate recent arguments that
intragenics are safer and more morally acceptable than transgenic
organisms, and more acceptable to the public, which might be thought to
justify a lower standard of regulation. We argue that the exemption of
intragenics from regulation is not justified, and that there may be
significant environmental risks associated with them. We conclude that
intragenics should be subject to the same standard of regulation as
other GMOs.

Document Number: 228

Via a historical reconstruction, this paper primarily demonstrates how
the societal debate on genetically modified organisms (GMOs) gradually
extended in terms of actors involved and concerns reflected. It is
argued that the implementation of recombinant DNA technology out of the
laboratory and into civil society entailed a "complex of concerns." In
this complex, distinctions between environmental, agricultural,
socio-economic, and ethical issues proved to be blurred. This fueled the
confusion between the wider debate on genetic modification and the risk
assessment of transgenic crops in the European Union. In this paper, the
lasting skeptical and/or ambivalent attitude of Europeans towards
agro-food biotechnology is interpreted as signaling an ongoing social
request - and even a quest - for an evaluation of biotechnology with
Sense and Sensibility. In this (re)quest, a broader-than-scientific
dimension is sought for that allows addressing the GMO debate in a more
"sensible" way, whilst making "sense" of the different stances taken in
it. Here, the restyling of the European regulatory frame on transgenic
agro-food products and of science communication models are discussed and
taken to be indicative of the (re)quest to move from a merely scientific
evaluation and risk-based policy towards a socially more robust
evaluation that takes the "non-scientific" concerns at stake in the GMO
debate seriously.

Document Number: 229

Biofortification of staple foods with iron (Fe) in the form of ferritin
(Ft) is now possible, both by conventional plant breeding methods and
transgenic approaches. Ft-Fe from plants and animals is absorbed well
(25-30%) by human subjects, but little is known about dietary factors
affecting its absorption. We used human intestinal Caco-2 cells and
compared Fe absorption from animal Ft and FeSO4 to determine the effects
of inhibitors and enhancers, such as phytic acid, ascorbic acid, tannic
acid, calcium and heme. When postconfluent cells were coincubated with
Fe-59-labeled (1 mu M) FeSO4 and dietary factors, at different molar
ratios of dietary factor to Fe (phytic acid:Fe, 10:1; ascorbic acid:Fe,
50:1; tannic acid:Fe, 50:1; calcium:Fe, 10:1 and hemin:Fe, 10:1), all
inhibited uptake from FeSO4, except ascorbate, confirming earlier
studies. In contrast, these dietary factors had little or no effect on
Fe uptake from undigested Ft or Ft digested in vitro at pH 4, except
tannins. However, results after in vitro digestion of Ft at pH 2 were
similar to those obtained for FeSO4. These results suggest that Fe
uptake occurs from both undigested as well as digested Ft but, possibly,
via different mechanisms. The Fe-Ft stability shown here could minimize
Fe-induced oxidation of Fe-supplemented food products. (c) 2008 Elsevier
Inc. 

Document Number: 230

Plants bind solar energy to organic matter via photosynthesis and
assimilation of carbon dioxide from the atmosphere and comprise the
major source of nutrition and bioenergy. Plant biotechnology contributes
to solution of important constraints in food and feed production and
creates new technologies and applications for the sustainable use of
plant resources. Genome-wide approaches such as massive parallel
sequencing and microarrays to study gene expression, molecular markers
for selection of important traits in breeding, characterization of
genetic diversity with the aforementioned approaches, and somatic
hybridization and genetic trans form at ion are important tools in plant
biotechnology. In this paper, studies carried out on enhanced resistance
to viruses and tolerance of cold stress in potato, genetic modification
of flower pigmentation and morphology in gerbera, production of edible
vaccines in transgenic barley seeds, and expression of heterologous
proteins for pharmaceutical purposes from vector viruses were chosen to
exemplify the general utility of biotechnological approaches and also
how plant biotechnology research has developed on cultivated plants at
University of Helsinki. The studies reveal cellular and genetic
mechanisms and provide scientific information that can be used for
widening the uses of crop plants. They can also be used to detect any
putative risks associated with the use of the biotechnological
application in agriculture and horticulture and to develop practises
which reduce any inadvertent negative consequences that plant production
may have to the environment.

Document Number: 231

Methods of analysis for products of modem biotechnology are required for
national and international trade in seeds, grain and food in order to
meet the labeling or import/export requirements of different nations and
trading blocks. Although many methods were developed by the originators
of transgenic events, governments, universities, and testing
laboratories, trade is less complicated if there exists a set of
international consensus-derived analytical standards. In any analytical
situation, multiple methods may exist for testing for the same analyte.
These methods may be supported by regional preferences and regulatory
requirements. However, tests need to be sensitive enough to determine
low levels of these traits in commodity grain for regulatory purposes
and also to indicate purity of seeds containing these traits. The
International Organization for Standardization (ISO) and its European
counterpart have worked to produce a suite of standards through open,
balanced and consensus-driven processes. Presently, these standards are
approaching the time for their first review. In fact, ISO 21572, the
"protein standard" has already been circulated for systematic review. In
order to expedite the review and revision of the nucleic acid standards
an ISO Technical Specification (ISO/TS 21098) was drafted to set the
criteria for the inclusion of precision data from collaborative studies
into the annexes of these standards.

Document Number: 232

Crop plants provide essential food nutrients to humans and livestock,
including carbohydrates, lipids, proteins, minerals and vitamins,
directly or indirectly. The level and composition of food nutrients vary
significantly in different food crops. As a result, plant foods are
often deficient in certain nutrient components. Relying on a single food
crop as source of nutrients thus will not achieve a balanced diet and
results in malnutrition and deficiency diseases, especially in the
developing countries, due mainly to poverty. The development and
application of biotechnology offers opportunities and novel
possibilities to enhance the nutritional quality of crops, particularly
when the necessary genetic variability is not available. While initial
emphasis of agricultural biotechnology has been placed on input traits
of crops such as herbicide tolerance, insect resistance and virus
resistance, increasing effort and promising proof-of-concept products
have been made in output traits including enhancing the nutritional
quality of crops since 1990s. Advancements in plant transformation and
transgene expression also allow the use of plants as bioreactors to
produce a variety of bio-products at large scale and low cost. Many
proof-of-concept plant-derived healthcare products have been generated
and several commercialized.

Document Number: 233

Sugar beet (Beta vulgaris) is an important arable crop, traditionally
used for sugar extraction, but more recently, for biofuel production. A
wide range of pests, including beet cyst nematode (Heterodera
schachtii), root-knot nematodes (Meloidogyne spp.), green peach aphids
(Myzus persicae) and beet root maggot (Tetanops myopaeformis), infest
the roots or leaves of sugar beet, which leads to yield loss directly or
through transmission of beet pathogens such as viruses. Conventional
pest control approaches based on chemical application have led to high
economic costs. Development of pest-resistant sugar beet varieties could
play an important role towards sustainable crop production while
minimising environmental impact. Intensive Beta germplasm screening has
been fruitful, and genetic lines resistant to nematodes, aphids and root
maggot have been identified and integrated into sugar beet breeding
programmes. A small number of genes responding to pest attack have been
cloned from sugar beet and wild Beta species. This trend will continue
towards a detailed understanding of the molecular mechanism of
insect-host plant interactions and host resistance. Molecular
biotechnological techniques have shown promise in developing transgenic
pest resistance varieties at an accelerated speed with high accuracy.
The use of transgenic technology is discussed with regard to
biodiversity and food safety.

Document Number: 234

There is a growing body of evidence suggesting that regular consumption
of foods rich in omega-3 long chain polyunsaturated fatty acids has
multiple positive health benefits. The fats and oils from marine fish
contain high contents of these beneficial fatty acids but increased
consumer demand has also increased strain on the ability of the world's
fisheries to meet demand from wild capture. Many consumers are choosing
fish oil supplements or are eating foods that have been complemented
with fish oils instead of consuming fish directly. However, removing
undesirable odors, flavors and contaminants is expensive. In contrast,
oils derived from land plants such as soybean are inexpensive and
contaminant free. Recent strides in plant molecular biology now allow
the engineering of oilseeds for the production of novel fats and oils,
including those synthesized by complex, multigene biosynthetic pathways
such as the omega-3 long chain polyunsaturated fatty acids. Given the
potential benefits to the environment with regards to overfishing and
the health prospects of increased consumption of these healthy fatty
acids, producing these fatty acids in oilseeds is a desirable and worthy
goal. In this review, we will describe the recent advances in this field
along with some of the technical hurdles encountered thus far.

Document Number: 235

Detection of genetically modified organisms in the food chain is an
important issue for all subjects involved in raw material control, food
industry and distribution. Both labeling and traceability of genetically
modified organisms are current issues that are considered for trade and
regulation. Currently, labeling of genetically modified foods containing
detectable transgenic material is required by the Brazilian legislation.
The Brazilian government published the Decree n degrees 4.680 in April
2003, which requires labeling for all foods or food ingredients, with a
stricter labeling threshold of M Although polymerase chain reaction
technology has some limitations, the high sensitivity and specificity
explain why it has been the first choice of most analytical laboratories
interested in detection of genetically modified organisms and their
derived products. Among the currently available methods, polymerase
chain reaction-based methods are accepted, considering the sensitivity
and reliability for detection of genetically modified-derived material
in routine analysis. In this paper a review of currently available
polymerase chain reaction methods for screening and quantifying
genetically modified-derived ingredients is presented, discussing their
applicability and limitations.

Document Number: 236

Transgenic lettuce (Lactuca sativa L. cv. 'Cortina') lines expressing
the asparagine synthetase A gene from Escherichia coli were produced to
alter the plant nitrogen status and eventually enhance growth. The
relative molecular abundance of water-soluble metabolites was measured
by H-1 NMR in transgenic and conventional plants at early developmental
stages and grown under the same conditions. NMR metabolic profiles
assessed that a transgenic, line and the wild-type counterpart shared
the same compounds, but it also revealed side effects on the carbon
metabolism following genetic modification. Concerning the nitrogen
status, the amino acid content did not vary significantly, except for
glutamic acid and gamma-aminobutyric acid, which diminished in the
transgenics. As for the carbon metabolism, in transgenic leaves the
contents of sucrose, glucose, and fructose decreased, whereas that of
inulin increased up to 30 times, accompanied by the alteration of most
Krebs's cycle organic acids and the rise of tartaric acid compared to
nontransformed controls. Lettuce leaf inulins consisted of short
oligomeric chains made of one glucose unit bound to two/four fructose
units. Inulins are beneficial for human health, and they are extracted
from plants and commercialized as long-chain types, whereas the short
forms are synthesized chemically. Hence, lettuce genotypes with high
content of foliar short-chain inulin represent useful materials for
breeding strategies and a potential source for low molecular weight
inulin.

Document Number: 237

The nutrients in the human diet ultimately come from plants. However,
all our major food crops lack certain essential vitamins and minerals.
Although a varied diet provides adequate nutrition, much of the human
population, particularly in developing countries, relies on staple
crops, such as rice or maize, which does not provide the full complement
of essential nutrients. Malnutrition is a significant public health
issue in most of the developing world. One way to address this problem
is through the enhancement of staple crops to increase their essential
nutrient content. Here, we review the current strategies for the
biofortification of crops, including mineral fertilization and
conventional breeding but focusing on transgenic approaches which offer
the most rapid way to develop high-nutrient commercial cultivars.

Document Number: 238

Metallothioneins (MTs), as a family of low-molecular-weight,
cysteine-rich, and metal-binding proteins, show potential for
utilization in functional food. Tomato plants were transformed with gene
constructs that contained mt-l encoding the mouse MT-l, similar in sense
orientation with the constitutively active double 35S promoter from
cauliflower mosaic virus. Three independent transformants, which had
copies of the gene in their genomes, were obtained. In these transgenic
lines, high-level expression of MT-l, high zinc content, and some
antioxidant enzyme activities were detected in leaves. The average zinc
content in transgenic tomato leaves was 32.7 mg/100 g FW, which about
1.6 times higher than that in wild-type. The superoxide dismutase
activity was also higher (68.6, 66.9, and 66.1 U/g FW in the three
transformants) than that in wild-type (57.4 U/g FW). In particular, the
levels of superoxide free radical scanvenging in the three transformants
were 14.2%, 14.6%, and 13.7%, respectively, which about 1.5 times higher
than that in control (5.6%). Transgenic MT tomato may potentially be
used as an antioxidant and for zinc supplementation.

Document Number: 239

The volatile compounds, 2-phenylacetaldehyde and 2-phenylethanol, are
important for the aroma and flavor of many foods, such as ripe tomato
fruits, and are also major constituents of scent of many flowers, most
notably roses. While much work has gone into elucidating the pathway for
2-phenylethanol synthesis in bacteria and yeast, the pathways for
synthesis in plants are not well characterized. We have identified two
tomato enzymes (LePAR1 and LePAR2) that catalyze the conversion of
2-phenylacetaldehyde to 2-phenylethanol: LePAR1, a member of the large
and diverse short-chain dehydrogenase/reductase family, strongly prefers
2-phenylacetaldehyde to its shorter and longer homologues (benzaldehyde
and cirinamaldehyde, respectively) and does not catalyze the reverse
reaction at a measurable rate; LePAR2, however, has similar affinity for
21-phenylacetaldehyde, benzaldehyde and cinnamaldehyde. To confirm the
activity of these enzymes in vivo, LePAR1 and LePAR2 cDNAs were
individually expressed constitutively in petunia. While wild type
petunia flowers emit relatively high levels of 2-phenylacetaldehyde and
lower levels of 2-phenylethanol, flowers from the transgenic plants
expressing LePAR1 or LePAR2 had significantly higher levels of
2-phenylethanol and lower levels of 2-phenylacetaldehyde. The in vivo
alteration of volatile emissions is an important step toward altering
aroma volatiles in plants. (C) 2007 Elsevier Ltd. 

Document Number: 240

Plant parasitic nematodes cause significant damage to crops on a
worldwide scale. These nematodes are often soil dwelling but rely on
plants for food and to sustain them during reproduction. Complex
interactions occur between plants and nematodes during the nematode life
cycle with plant roots developing specialized feeding structures through
which nematodes withdraw nutrients. Here we describe a novel method for
delivering macromolecules to feeding nematodes using a virus-based
vector [tobacco rattle virus (TRV)]. We show that the parasitic nematode
Heterodera schachtii will ingest fluorescent proteins transiently
expressed in plant roots infected with a TRV construct carrying the
appropriate protein sequence. A prerequisite for this delivery is the
presence of replicating virus in root tips prior to the formation of
nematode-induced syncytia. We show also that TRV vectors expressing
nematode gene sequences can be used to induce RNAi in the feeding
nematodes.

Document Number: 241

Genetically modified Bacillus thuringiensis Berliner (Bt) maize (Zea
mays L.) expressing Cry toxins against various target pests is now grown
on more than 16 million hectares worldwide, but its potential effects on
the soil ecosystem need to be further investigated. In an 8-month field
study, we investigated the effects of Bt maize expressing the Cry1Ab
protein on both the soil community and maize residue decomposition. We
used litterbags with three different mesh sizes (20, 125, and 5,000 mu
m) to investigate potential effects of different soil organism groups on
the decomposition processes. Litterbags were incorporated into the soil
in fall into a field that had previously been planted with non-Bt maize
and subsamples were removed monthly. The dry weight of the remaining
residue was measured for all bags. Bt and non-Bt maize decomposed
similarly in large mesh bags, which allowed the whole soil organism
community to enter and interact with each other. In contrast, Bt maize
decomposed significantly faster than non-Bt maize at some sample dates
in winter in bags with small and medium mesh sizes. At the end of the
experiment in late spring, however, there was no significant difference
in the amount of maize plant residues remaining for any of these three
mesh sizes. Additionally, soil organisms from bags with the largest mesh
size were identified. The most frequent taxa extracted were collembolans
(Isotomidae, Tullbergiidae, Entomobryidae), mites (Gamasina, Oribatida),
and annelids (Enchytraeidae). Three of these taxa were extracted in
higher numbers from non-Bt than Bt residue (Tullbergiidae, Gamasina,
Enchytraeidae), while there was no difference in the number of
individuals extracted for the remaining three taxa. Our results do not
show major changes in the decomposition of Bt maize residue and in the
composition of the soil organism community. However, further studies are
needed that assess the impact of the continuous release of Cry1Ab via
root exudates and plant biomass on the soil ecosystem.

Document Number: 242

In this paper, four different physical treatments (microwaves, heating
by conduction, sonication and pressure autoclaving) were performed to
degrade a pure DNA extract, and their influence on GMO quantification
was studied. The aim was to check the hypothesis that processing of
agrofood products results in a similar degradation rate for both the
transgenic target and the specific target. Indeed we could observe that
even if the used physical treatments could lead to a significant
increase of C-t values for both transgenic and specific targets, the
resulting Delta C-t remained stable. So, the main conclusion of the
study is that the aforementioned hypothesis seems valid and thus a
physical degradation of DNA will not affect the relative quantitation of
the GMO content, provided that both the specific and transgenic targets
have very similar size. A second important issue of the experiments
performed was that DNA is a very robust molecule as it is extremely
difficult to reach a mean size below 100 bp. The study also gives
evidence of the importance of using small targets.

Document Number: 243

The remarkable amount of public and scientific debate with regard to the
safety of genetically modified (GM) crops has resulted in the
implementation of mandatory risk assessments of newly developed GM
crops, as well as mandatory labeling of GM crops and foods following
their commercial release. Among the many currently available GM rice
lines, transgenic rice lines that overexpress a trehalose6-phosphate
synthase/phosphatase (TPSP) fusion gene have attracted particular
interest from researchers, by virtue of their superior tolerance to a
variety of abiotic stresses. The primary objective of this study was te.
determine the copy numbers of the transgene element, and to identify the
genomic sequences flanking the integration site of the transgene element
in a selected TPSP transgenic rice line, in order to develop new
event-specific detection techniques. The genomic sequences flanking the
integration site of the transgene element were identified via an inverse
PCR protocol. It was determined that a single copy of the TPSP fusion
had been integrated into the transgenic line. An effort to find a rice
sequence usable as an internal positive control for the screening of the
transgenic rice resulted in the identification of an RBE4 gene sequence,
which appears as one copy within the rice genome, and is rice-specific.
The characterization of the genomic sequences flanking the transgene, as
well as the availability of the internal positive control sequence,
enabled the design of a qualitative PCR technique and a quantitative
TaqMan-based detection method for this transgenic rice line. The results
of this study may prove useful with regard to the large-scale screening
of this transgenic rice line in the processes of risk assessment and
commercialization, as well as in the development of novel methods for
the detection of other transgenic rice lines. (c) 2006 Elsevier Ltd. All
rights reserved.

Document Number: 244

Despite being introduced more than a decade ago, agricultural
biotechnology still remains framed in controversy impacting both the
global economy and international regulations. Controversies surrounding
agricultural biotechnology produced crops and foods commonly focus on
human and environmental safety, intellectual property rights, consumer
choice, ethics, food security, poverty reduction and environmental
conservation. Originally, some consumers were reluctant to accept the
first generation agricultural biotechnology products because they
appeared to primarily benefit agricultural producers; however, it is
clear from continued evaluations that these technologies also improved
both the safety and wholesomeness of food and helped improve the
environment. Plants engineered to resist insect pests and tolerate less
toxic pesticides resulted in improved yields thereby enabling farmers to
produce more food per acre while reducing the need for herbicides,
pesticides, and water and tilling. An indirect benefit of reduced pest
damage in transgenic corn expressing genes to control insect pests is
lower levels of mycotoxins, most notably those caused by the genus
Fusarium. Mycotoxins are an important regulatory issue globally because
of their toxic and carcinogenic potential to humans and animals.
Complicating this issue is the fact that toxicological databases for
mycotoxins are relatively incomplete compared to other food
contaminants. Current debates about agricultural biotechnology and
mycotoxins reveal significant differences in perception of associated
risks and benefits. When faced with uncertainty, regulators tend to set
limits as low as possible. Additionally, some regulators invoke the ''
Precautionary Principle '' when limited information is available or
disputes over interpretation exist for possible contaminants, including
mycotoxins. A major concern regarding use of the '' Precautionary
Principle '' is the appearance that regulators can justify setting any
limit on the basis of inconclusive or unknown potential hazards of a
contaminant which may significantly impact global trade because
mycotoxin residues vary widely between countries. This paper describes
the current economic and heath impact of these regulations and their
impact on international trade. Published by Elsevier B.V.

Document Number: 245

Except for the events RT73, MS8, RF3, and T45, event-specific detection
methods for most commercialized genetically modified (GM) rapeseed
varieties have not been established, and as a result, the enforcement of
genetically modified organism labeling policies has been hindered. The
genetically modified rapeseeds, MS1 x RF1 and MS1 x RF2, are 2 of 11
approved GM-rapeseed varieties for commercialization. In this study, the
right border junction fragments between the gene construct and the
rapeseed genome of events RF1, RF2, and MS1 were isolated using the
commercially available GenomeWalker technology. Homology analysis
indicated that the gene construct of RF1 integrated upstream of the
nuclease gene, and that of the RF2 and MS1 inserted into the exon region
of a gene encoding for an unknown protein. The event-specific primer
pairs and corresponding probes were designed on the basis of the
revealed right border junction fragments. Then, we successfully
developed the identification and quantification methods for the
gene-stacked hybrids MS1 x RF1 and MS1 x RF2 using those primers and
probes. The relative limit of detection in the qualitative polymerase
chain reaction (PCR) was 0.013% for the RF2 and MS1 assays using 100 ng
of rapeseed DNA per reaction and 0.13% for the RF1 assay. The absolute
limit of detection in the quantitative PCR was approximately one to two
initial copies for each of the three event-specific assays. The
evaluation of the real-time PCR assays revealed that the qualitative and
quantitative methods developed by focusing on the gene-stacked hybrids
MS1 x RF1 and MS1 x RF2 were highly specific, sensitive, and suitable
for samples with a low quantity of DNA.

Document Number: 246

Improvements in production methods over the last two decades have
resulted in aquaculture becoming a significant contributor to food
production in many countries. Increased efficiency and production levels
are off-setting unsustainable capture fishing practices and contributing
to food security, particularly in a number of developing countries. The
challenge for the rapidly growing aquaculture industry is to develop and
apply technologies that ensure sustainable production methods that will
reduce environmental damage, increase productivity across the sector,
and respect the diverse social and cultural dimensions of fish farming
that are observed globally. The aquaculture industry currently faces a
number of technology trajectories, which include the option to
commercially produce genetically modified (GM) fish. The use of genetic
modification in aquaculture has the potential to contribute to increased
food security and is claimed to be the next logical step for the
industry. However, the potential use of these technologies raises a
number of important ethical questions. Using an ethical framework, the
Ethical Matrix, this paper explores a number of the ethical issues
potentially raised by the use of GM technologies in aquaculture. Several
key issues have been identified. These include aspects of distributive
justice for producers; use of a precautionary approach in the management
of environmental risk and food safety; and impacts on the welfare and
intrinsic value of the fish. There is a need to conduct a comparative
analysis of the full economic cycle of the use of GM fish in aquaculture
production for developing countries. There is also a need to initiate an
informed dialogue between stakeholders and strenuous efforts should be
made to ensure the participation of producers and their representatives
from developing nations. An additional concern is that any national
licensing of the first generation of GM fish, i.e., in the USA, may
initiate and frame an assessment cycle, mediated by the WTO, which could
dominate the conditions under which the technology will be applied and
regulated globally. Therefore, an integrated analysis of the technology
development trajectories, in terms of international policy, IPR, and
operational implications, as well as an analysis of a broader range of
ethical concerns, is needed.

Document Number: 247

For cassava to become a safe and acceptable crop, it is necessary to
reduce the cyanogen levels in cassava foods. While this objective can be
achieved by processing procedures, recent findings have shown that it is
also possible to achieve it by suppression of cyanogen synthesis or by
accelerating cyanogen turnover and volatilization. In 2003,
cyanogen-free cultivars were generated by selective inhibition
CYP79D1/D2 gene expression. The CYP79D1/D2 enzymes catalyze the
first-dedicated step in cyanogen synthesis. Tissue-specific inhibition
of CYP79D1/D2 expression in leaves lead to a 99% reduction in root
cyanogen levels, indicating that the cyanogenic glycoside, linamarin, is
synthesized in leaves and transported to roots. An alternative strategy
to the reduce cyanogen content is to enhance cyanogen detoxification and
cyanide volatilization during processing. This strategy has the
advantage that cyanogen levels in unprocessed roots are not altered,
potentially providing protection against herbivory and/or theft. To
produce cultivars that promote rapid cyanide volatilization,
hydroxynitrile lyase (HNL), which catalyzes the last step in
cyanogenesis, was overexpressed in roots. Elevated HNL activity resulted
in a 3-fold increase in the rate of cyanogen turnover. Importantly, the
cyanogen content of the transformed and wild-type plants was identical,
a potential benefit for farmers.

Document Number: 248

Agriculture plays an important role in the national economy of Pakistan,
where most of the rapidly increasing population resides in rural areas
and depends on agriculture for subsistence. Biotechnology has
considerable potential for promoting the efficiency of crop improvement,
food production, and poverty reduction. Use of modern biotechnology
started in Pakistan since 1985. Currently, there are 29 biotech
centers/institutes in the country. However, few centers have appropriate
physical facilities and trained manpower to develop genetically modified
(GM) crops. Most of the activities have been on rice and cotton, which
are among the top 5 crops of Pakistan. Biotic (virus/bacterial/insect)
and abiotic (salt) resistant and quality (male sterility) genes have
already been incorporated in some crop plants. Despite acquiring
capacity to produce transgenic plants, no GM crops, either produced
locally or imported, have been released in the country. Pakistan is
signatory to the World Trade Organization, Convention on Biological
Diversity, and Cartagena protocols. Several legislations under the
Agreement on Trade-Related Aspects of Intellectual Property Rights have
been promulgated in the country. National Biosafety Guidelines have been
promulgated in April 2005. The Plant Breeders Rights Act, Amendment in
Seed Act-1976, and Geographical Indication for Goods are still passing
through discussion, evaluation, and analysis phases. Meanwhile, an
illegal GM crop (cotton) has already sneaked into farmer's field.
Concerted and coordinated efforts are needed among various ministries
for implementation of regulation and capacity building for import/export
and local handling of GM crops. Pakistan could easily benefit from the
experience of Asian countries, especially China and India, where
conditions are similar and the agriculture sector is almost like that of
Pakistan. Thus, the exchange of information and experiences is important
among these nations.

Document Number: 249

The present study investigates the variation of Fe concentration and
partitioning in different grain tissues, particularly in the endosperm
(white rice) among different rice cultivars from diverse genetic
backgrounds. Iron concentration ranged from 10 to 20 mgkg(-1) in brown
rice and 3-11 mgkg(-1) in white rice. Iron concentration in white rice
was not correlated with that in brown rice. Polishing removed 25-84% of
Fe from the brown rice, resulting in a low Fe concentration in the white
rice and changed the ranking order for Fe concentration among the
cultivars tested. Total Fe content in the whole grain including husk (r
= 0.28, NS) and total Fe content in brown rice (r = 0.09, NS) was not
significantly correlated with Fe concentration in white rice. Iron
content in white rice was found to vary between 0.05 and 0.2 mu g
grain(-1) among cultivars tested, with the highest Fe content in cv.
Sakha. Iron concentration in white rice was negatively correlated with
total Fe content in the bran fraction, which is lost during the
polishing process to produce white rice from brown rice (r = -0.82**).
The partitioning of Fe between the endosperm and bran seemed to play a
key role in achieving high Fe concentration in the white rice. The
surface cell layers of the caryopsis appeared to be the main resistance
of Fe partitioning into the endosperm (white rice) as there was a
significantly negative correlation between Fe concentration in the white
rice and Fe content in the bran, which is lost during the polishing
process. (C) 2007 Society of Chemical Industry

Document Number: 250

Using 2.5% (w/w) transgenic Roundup Ready soybean as raw material, the
effects of critical processing procedures on the fragment sizes of
endogenous gene lectin and transgenic gene epsps and the quantity of
transgenic component during bean curd preparation were analysed. The
results showed that the different processing procedures differed in
their impact on the degradation of endogenous and transgenic genes and
that the transgenic content of the intermediate products and final
product was consequently changed. Physical mechanical action such as
jordaning caused both endogenous and transgenic genes to degrade but had
more effect on the latter. The jordaning process degraded the DNA of
endogenous gene from 1883 to 836 bp and that of transgenic gene from
1512 to 408 bp. The adding CaSO4 process, predominantly a chemical
reaction, had less influence on the transgenic gene than on the
endogenous gene, the latter being degraded from 836 to 407 bp. The
extrusion moulding process resulted in a significant degradation of DNA
in transgenic gene from 408 to 190 bp but had no obvious effect on the
endogenous gene. After the critical processing procedures of jordaning,
syruping, adding CaSO4 and extrusion moulding during bean curd
preparation the transgenic content of the intermediate products and
final product was 1.656, 0.435, 1.150 and 0.797% respectively. (c) 2007
Society of Chemical Industry.

Document Number: 251

Nutritional assessment of transgenic crops used for human food and
animal feed is an important aspect of safety evaluations. A
insect-resistant rice (IRR) was generated by the stable insertion of
sck, a modified cow-pea trypsin inhibitor gene, and cry1Ac encoding a
crystal protein from Bacillus thuringiensis into the genome of a common
variety of Chinese indica rice. The composition of the brown and milled
rice grain from the resulted IRR line designated Liangyou Kefeng No. 6
was compared with that of the parental rice cultivar Liangyou 2186.
Nutrients, including the proximates, amino acids, fatty acids, minerals,
and vitamins, were measured. The antinutritive components such as phytic
acid, lectin, and trypsin inhibitors were also examined. The data
demonstrated that the nutritional quality of both the brown and milled
rice grains from the transgenic line was substantially equivalent to
values reported for other commercial lines.

Document Number: 252

Food and Agriculture Organization/World Health Organization (FAO/WHO)
recommended that IgE cross-reactivity between a transgenic protein and
allergen be considered when there is >= 35% identity over a sliding
"Window" of 80 amino acids. Our objective was to evaluate the false
positive and negative rates observed using the FAO/WHO versus
conventional FASTA analyses. Data used as queries against allergen
databases and analyzed to assess false positive rates included: 1102
hypothetical corn ORFs; 907 randomly selected proteins; 89 randomly
selected corn proteins; and 97 corn seed proteins. To evaluate false
negative rates of both methods: Bet v la along with several
crossreacting fruit/veaetable allergens and a bean a-amylase inhibitor
were used as queries. Both methods were also evaluated for their ability
to detect a putative nonallergenic test protein containing a sequence
derived from Ara h 1. FASTA versions 3.3t0 and 3.4t25 were utilized.
Data indicate a conventional FASTA analysis produced fewer false
positives and equivalent false negative rates. Conventional FASTA versus
sliding window derived E scores were generally more significant. Results
suggest a conventional FASTA search provides more relevant identity to
the query protein and better reflects the functional similarities
between proteins. It is recommended that the conventional FASTA analysis
be conducted to compare identities of proteins to allergens.

Document Number: 253

Many important health promoting and functional characteristics are
attributed to the non-digestible polysaccharide, inulin. Its use as a
prebiotic in functional food and feed has further increased inulin
demand worldwide. Inulin production in crops used for food and feed
application, such as maize and potato, may provide a more direct and
cost-effective route to provide functional food or feed when compared to
native inulin producers. Towards this end we have expressed the inulin
synthesizing enzymes, I-SST and I-FFF from Jerusalem artichoke in maize
and potato. Transgenic maize plants produced inulin type fructan (at 3.2
mg/g kernel) when expressing an endosperm targeted gene cassette. Kernel
development and germination were not affected. Potato tubers expressing
1-sst accumulated 1.8 mg inulin/g tuber while tubers with a combined
expression of 1-sst and 1-fft accumulated 2.6 mg inulin/g tuber. Inulin
accumulation in maize kernels was modulated by kernel development.
Inulin levels peaked and then underwent moderate degradation by late
kernel development. In potato, inulin production was relatively stable
throughout tuber development and little evidence of degradation was
observed. The accumulation of 1-kestose in transgenic maize was
positively correlated with kernel sucrose concentration. Introduction of
the fructan synthetic pathway in a high-sucrose maize background
increased inulin accumulation to 41 mg/g kernel. Evidence is presented
indicating that sucrose availability is limiting fructan production in
transgenic maize. (c) 2007 Elsevier Ireland Ltd. 

Document Number: 254

Rice is the most important staple food in the world. The rapid
development of transgenic rice and its future commercialization have
raised concerns regarding transgene flow and its potential environmental
risk. It is known that rice is a self-pollinated crop; the outcrossing
rate between common cultivars is generally less than 1%. In order to
improve the detection sensitivity of rice transgene flow, a male sterile
(ms) line BoA with a high outcrossing rate was used as a pollen detector
in this study. A concentric circle design was adopted, in which the
transgenic rice B2 containing bar gene as a pollen donor was planted in
the center circle and the recipient BoA was planted in eight compass
sectors. The frequency of transgene flow in compass sectors was analyzed
by continuous sampling to generate cumulative data. The results of two
years with sound reproducibility demonstrated that the rice gene flow
was closely associated with the wind direction. According to the mean
frequency of transgene flow, the eight sectors can be divided into two
groups: a higher frequency group downstream of the prevailing wind (DPW)
with a mean frequency ranging from 6.47 to 26.24%, and a lower frequency
group lateral to or upstream of the prevailing wind (UPW) with a mean
frequency of 0.39 to 3.03%. On the basis of the cumulative data, 90-96%
of the cumulative gene flow events occurred in the four DPW sectors,
while it was 4-10% in the four UPW sectors. By using these systematic
data, simulation models and isograms of transgene flow in the eight
compass sectors were calculated and drawn, respectively.

Document Number: 255

Genomics is providing a new set of tools for cereal chemistry. Analysis
of the DNA of the major cereals is increasing the understanding of the
basis of grain quality at the gene level. Differences in the sequence of
the gene in different cultivars or differences in the level of
expression of the gene may be used to explain differences in processing
or end-use quality characteristics. For example, quantitative analysis
of the levels of expression of genes at different stages during seed
development in wheat or germination in barley can be used to define the
genetic basis of differences in wheat and barley quality. Rice quality
traits such as fragrance and gelatinization temperature (cooking
temperature) can be explained by DNA sequence differences in specific
genes identified using genomics approaches. Rapid and reliable species
and cultivar identification based on DNA analysis methods developed
using genomics tools can be applied to grain and to food products. This
technology has special advantages in the analysis of complex mixtures of
cereals. Technologies for very high-throughput and very low-cost
analysis of large numbers of samples are now available. These may be
applied by cereal chemists at many levels: selection in cereal breeding,
optimizing processing, and analysis of the identity and composition of
grain or food samples.

Document Number: 256

On a worldwide basis, parasitic weeds represent one of the most
destructive and intractable problems to agricultural production in both
developed and developing countries. About 20 families (3,000-5,000
species) of higher plants are parasitic on the plant kingdom and may
cause production losses of 30-80% in staple food and industrial crops on
every continent. Compared with the other weeds, parasitic weeds are
difficult to control by conventional means because of their life style:
Parasites are intimately involved with the host and have so much
metabolic overlap with the host that differential treatments are very
difficult to develop. In some cases, the parasites are closely
associated to the host root, concealed underground, and undiagnosed
until they irreversible damage the crop. Several different approaches
(cultural, mechanical, chemical, use of resistant varieties, and
biological) to control parasitic weeds are currently in use, but are
only partially successful. Recent reviews have covered the physiology
and interactions between parasitic plants and their hosts, taxonomy, and
the biology and classical control of parasitic weeds. The current review
will discuss why alternative methods are needed to control parasitic
weeds and will summarize conventional and new biotechnology-based
control measures against the major world pests Striga, Orobanche,
Cuscuta, and mistletoes (Phoradendron and Viscum genera). Effectiveness,
advantages and disadvantages, environment safety, and simplicity of
these new biotechnological methods will be reviewed.

Document Number: 257

China is a developing country with the biggest population in the world.
Whether the problem in grain security of China can be properly solved
has the direct bearing on the order of the world grain market. On the
basis of experience data from 1978 to 2003 and the prediction of future
demand, we got the conclusion that the food security poses a threat to
China and the development and extensive application of agricultural
biotechnologies may solve this problem, Transgenic technology may become
the agricultural biotechnology that possesses independent intelligent
property and leading position across the world.

Document Number: 258

Nutritional genomics is a new and promising science area which can
broadly be defined as the application of high throughput genomics
(transcriptomics, proteornics, metabolomics/metabonomics) and functional
genomic technologies to the study of nutritional sciences and food
technology. First utilised in the food industry by plant
biotechnologists to manipulate plant biosynthetic pathways, the use of
genomic technologies has now spread within the agriculture sector,
unleashing a host of new applications (e.g. approaches for producing
novel, non-transgenic plant varietals; identification of genetic markers
to guide plant and animal breeding programmes; exploration of diet-gene
interactions for enhancing product quality and plant/animal health).
Beyond agriculture, genomic technologies are also contributing to the
improvement of food processing, food safety and quality assurance as
well as the development of functional food products and the evolution of
new health management concepts such as 'personalised nutrition', an
emerging paradigm in which the diet of an individual is customised,
based on their own genomic information, to optimise health and prevent
disease. In this review the relevance of nutritional genomics to the
food industry will be considered and examples given on how this science
area is starting to be leveraged for economic benefits and to improve
human nutrition and health.

Document Number: 259

The article is to establish multiplex PCR method for quantitative
detecting transgenic component promoter derived from Cauliflower Mosaic
Virus (CaMV 35S) and nos terminator derived from Agrobacterium
tumefaciens (Tnos) in foods. According to the specific sequence of CaMV
35S and Tnos which have been used in genetically modified organisms
(GMOs) frequently, and the sequence of soybean endogenous lectin gene,
three pairs of primers and corresponding fluorophore double stranded
probes (FDSP) were designed to allow for quantitative detecting of GMOs.
FDSP designed with maximal specificity also showed the greatest
detection sensitivity, and the ease in design, the simple single-dye
labeling chemistry. FDSP-multiplex quantitative PCR (FDSP-MQPCR) methods
were established for the detection of transgenic component CaMV 35S and
Tnos simultaneously. Ten soybean flour samples were tested with
FDSP-MQPCR method. The method gives five positive-samples with
quantitative results in 5 h, and accuracy rate is above 97.0%. The
described methods enabled a sensitive, specific, simple, and accurate
detection of transgenic component and thus provide a useful tool for
quantitative analysis of raw and processed food products. FDSP-MQPCR
method has not only improved detection efficiency and result
credibility, but also has guaranteed the better accuracy and
repetitiveness. (c) 2005 Published by Elsevier Ltd.

Document Number: 260

Ferritin is present in several types of plants in low, concentrations,
but it is possible to enhance this content by plant breeding, or by
inserting the gene for ferritin into staple foods I. Since each ferritin
molecule can bind thousands of iron atoms, this may be a, sustainable
means to increase the iron content of plants. Before launching such
efforts it is important to determine whether ferritin-bound iron is
bioavailable. We assessed this in vitro using Caco-2 cells and in vivo
using radiolabeled ferritin and whole body counting in human subjects.
In Caco-2 cells, we found that dietary factors affecting iron
absorption, such as ascorbic acid, phytate, and calcium, had very
limited effect on iron uptake from intact ferritin, suggesting that
ferritin-bound iron is absorbed via a mechanism different from that of
non-heme iron. Using in vitro digestion, we found that ferritin was
relatively resistant against proteolytic enzymes. Binding of ferritin to
Caco-2 cells was found to be saturable and the kinetics for binding
characteristic for a receptor-mediated process. In human subjects; we
found that iron ab sorption from animal ferritin was similar to that
from ferrous sulfate, suggesting that iron is well absorbed from
ferritin. We did not find any significant difference between iron
absorption from ferritin reconstituted with high-phosphate (plant-type)
and low-phosphate (animal-type) ferriltin mineral, suggesting that plant
ferritin-iron is bioavailable. In a subsequent human study we also found
that iron from purified soybean ferritin given in a meal was as well
absorbed as ferrous iron. In conclusion, iron is well absorbed from
phytoferritin and may represent a means of biofortification of staple
foods.

Document Number: 261

Genetically modified orgainsms and food produced of these are subject to
careful examination before they are put into circulation. If they are
granted non-restricted admission, one can be sure that the food does not
affect human health. The present contribution describes the various
steps of safety evaluation and its methodology Great emphasis is laid on
studies of the allergenic potential and On the discussion of markets of
antibiotic resistance. Novel proteins in this genetically modified food
do not represent a greater risk of food allergies than other food
proteins. The danger that antibiotic-resistance markets Used in
genetically modified food may lead to an additional dissemination of
these genes in bacteria is very low. It is desirable, however, that
alternative chances of selection be soon available to guarantee as much
safety as possible and to dispel frequently irrelevant doubts.

Document Number: 262

By taking 2.5% (w/w) transgenic Roundup Ready Soybean as raw materials,
with the detecting technology of traditional PCR and real-time
fluorescence quantitative PCR, this study analyzed the influence of such
critical processing procedures as jordaning, siruping, blending,
homogenization and sterilization in soymilk preparation on the fragment
size of endogenous gene lectin and exogenous gene epsps, as well as the
quantification of transgenic component in Roundup Ready Soybean, and
discussed the degradation mechanism of DNA in the course of transgenic
soybean processing. The results indicated that, different processing
procedures differ in their impact on the degradation of endogenous gene
and exogenous gene. Physical mechanical action, such as the jordaning,
can make endogenous and exogenous genes degrade, and has more effect on
the later. The jordaning process degraded the DNA of endogenous gene
from 1883 to about 836 bp, and exogenous gene from 1512 bp to about 408
bp. In the blending procedure, exogenous gene DNA was also degraded from
about 408 to 190 bp, but there was no obviously action on the endogenous
gene. After the endogenous and the exogenous genes were degraded to some
degree, such as 836 and 408 bp, respectively, they were not evidently
affected by siruping procedure at 100 degrees C for 15 min. However,
endogenous gene was further considerably degraded from around 836 to 162
bp in the sterilization procedure at 121 degrees C for 30 s. The effect
of the homogenization step on endogenous and exogenous genes was similar
to that of siruping procedure. In addition, jordaning and siruping
greatly damaged to exogenous gene, which rapidly decreased the
transgenic content from 2.5 to 1.656% and 0.435%, respectively. While
the endogenous gene was not evidently affected by the blending and
homogenization procedures, the exogenous gene DNA was remarkably
degraded in the two procedures. Therefore, the transgenic content
continued decreased to 0.377% after homogenization procedure. Hereafter,
the endogenous gene was further considerably degraded more than the
exogenous gene in the sterilization procedure, which caused a little
bounce of the transgenic content in the final product to 0.518%.

Document Number: 263

The mature coffee grain contains a high level of chlorogenic acids
(CGA). We have quantified the main caffeoylquinic acids (CQA) and
dicaffeoylquinic acids (diCQA) in Coffea canephora (robusta) grain
during late development. This analysis indicates the CQA levels remain
relatively steady during the final stages of grain development, but the
levels of diCQA fall significantly. Analysis of quinic acid, a key CGA
precursor in coffee, shows it is present at a high level in the early
grain, but then drops to a low level as development progresses. To
better understand coffee CGA synthesis, and to learn why diCQA and
quinic acid levels fall late in grain development, we have cloned cDNA
encoding four key enzymes for CGA synthesis in coffee; HCT, HQT, C3H1
and CCoAOMT1. The characterization of recombinant HCT, HQT and CCoAOMT1
proteins is also described. Quantitative real-time RT-PCR data is
presented for different stages of grain and pericarp development, as
well as several other C. canephora tissues. Elevated HCT and CCoAOMT1
expression in branch tissues strongly suggests the products of these
genes are associated with increased lignin synthesis, while higher HQT
expression appears to be more closely correlated with CGA accumulation.
The data presented forms an important base for designing new experiments
aimed at improving our understanding of CGA synthesis in coffee and
other plants, and could facilitate the development of new strategies to
increase the CGA content of plant foods. (c) 2007 Elsevier Ireland Ltd.

Document Number: 264

Expression of a growth hormone (GH) transgene can achieve dramatic
impacts upon the performance of aquaculture stocks, for example,
increasing the growth rate of coho salmon 11-fold. With the prospect of
improved production efficiency, some aquaculturists want to produce
transgenic fish commercially. Commercialization of transgenic fish and
shellfish, however, poses not only ecological, food safety, and
regulatory issues, but also animal welfare concerns. We review the
impacts of GH transgene expression on the behavior and welfare of
cultured fishes, focusing on Atlantic salmon, coho salmon, tilapia and
common carp lines that have been posed for commercial production.
Elevation of GH increases rates of protein synthesis and lipid
mobilization, affecting not only growth, but also feed conversion
efficiency, metabolic rate, body composition, head and body
morphometrics, osmoregulation, and age at maturity, with other
modifications particular to species and transgenic lines. Because of
heightened feeding motivation, transgenic fishes often prove more
active, aggressive, and willing to risk exposure to predation. Swimming
ability of some transgenic lines is reduced. While limited in scope,
studies suggest that fish of some transgenic lines show lessened
reproductive behavior. Welfare issues posed by morphological,
physiological, and behavioral alternation in GH-transgenic fishes have
not been well characterized. The use of weaker promoters in expression
vectors and by selection of transgenic lines with physiologically
appropriate levels of GH expression may reduce welfare issues that do
arise. Also, markers of welfare for GH-transgenic fish under production
conditions can be monitored to determine how culture systems and
practices can be modified to promote fish welfare. GH-transgenic fish
may be the first transgenic animals commercialized for food production;
with other transgenic fishes likely to become widely produced, a range
of behavioral and welfare issues will follow. (c) 2006 Elsevier B.V. All
rights reserved.

Document Number: 265

Background: Rice is both a food source for a majority of the world's
population and an important model system. Available functional genomics
resources include targeted insertion mutagenesis and transgenic tools.
While these can be powerful, a non- transgenic, unbiased targeted
mutagenesis method that can generate a range of allele types would add
considerably to the analysis of the rice genome. TILLING ( Targeting
Induced Local Lesions in Genomes), a general reverse genetic technique
that combines traditional mutagenesis with high throughput methods for
mutation discovery, is such a method.
Results: To apply TILLING to rice, we developed two mutagenized rice
populations. One population was developed by treatment with the chemical
mutagen ethyl methanesulphonate ( EMS), and the other with a combination
of sodium azide plus methyl- nitrosourea (Az- MNU). To find induced
mutations, target regions of 0.7 - 1.5 kilobases were PCR amplified
using gene specific primers labeled with fluorescent dyes.
Heteroduplexes were formed through denaturation and annealing of PCR
products, mismatches digested with a crude preparation of CEL I nuclease
and cleaved fragments visualized using denaturing polyacrylamide gel
electrophoresis. In 10 target genes screened, we identified 27
nucleotide changes in the EMS- treated population and 30 in the AzMNU
population.
Conclusion: We estimate that the density of induced mutations is two- to
threefold higher than previously reported rice populations ( about 1/300
kb). By comparison to other plants used in public TILLING services, we
conclude that the populations described here would be suitable for use
in a large scale TILLING project.

Document Number: 266

PCR products obtained by selective amplification of transgenic DNA
derived from food samples containing Roundup Ready soybean or Bt-176
maize have been analyzed by anion-exchange HPLC. Peptide nucleic acids
(PNAs), oligonucleotide analogues known to bind to complementary
single-stranded DNA with high affinity and specificity, have been used
as specific probes in order to assess the identity of the peaks
observed. Two different protocols were adopted in order to obtain
single-stranded DNA: amplification with an excess of one primer or
digestion of one DNA strand. The single-stranded DNA was mixed with the
PNA probe, and the presence of a specific sequence was revealed through
detection of the corresponding PNA:DNA peak with significantly different
retention time. Advantages and limits of this approach are discussed.
The method was tested with reference materials and subsequently applied
to commercial samples.

Document Number: 267

Crops are being genetically engineered to produce a wide variety of
drugs, vaccines and other pharmaceutical proteins. Although these crops
may open the door to less expensive and more-readily available drugs,
there is concern regarding the potential for contamination of human food
and livestock feed, as well as environmental harm. The outlook for the
production of pharmaceutical crops in California currently appears mixed
To date, 18 federal permits for field trials involving pharmaceutical or
industrial proteins have been approved in California. However, the
state's farming community and general public have thus far rejected
pharmaceutical crop production, and a handful of local governments have
recently banned the cultivation of genetically modified crops, including
pharmaceutical crops. In light of the many pros and cons, three major
approaches - the precautionary approach, risk analysis and cost-benefit
analysis - could be used to move the debate about pharmaceutical crops
forward.

Document Number: 268

Numerous clinical studies have demonstrated the cardiovascular and
mental health benefits of including very long chain omega-3
polyunsaturated fatty acids, namely eicospentaenoic acid (EPA) and
docosohexaenoic acid (DHA) in the human diet. Certain fish oils can be a
rich source of omega-3 long chain polyunsaturated fatty acids although
processed marine oils are generally undesirable as food ingredients
because of the associated objectionable flavors and contaminants that
are difficult and cost-prohibitive to remove. Oilseed plants rich in
omega-3 fatty acids, such as flax and walnut oils, contain only the
18-carbon omega-3 polyunsaturated fatty acid alpha-linolenic acid, which
is poorly converted by the human body to EPA and DHA. It is now possible
to engineer common omega-6 rich oilseeds such as soybean and canola to
produce EPA and DHA and this has been the focus of a number of academic
and industrial research groups. Recent advances and future prospects in
the production of EPA and DHA in oilseed crops are discussed here.

Document Number: 269

Salinity stress is one of the most serious factors limiting plant growth
and threatening global food security. Identification of salt tolerance
relevant genes from halophytes can provide more insights into the
determinants of salt stress tolerance in plants. In this study,
functional screening of genes for the salinity tolerance in
Thellungiella halophila (salt cress) was carried out using fission yeast
as a host organism. A cDNA library of the salt cress, constructed in the
yeast vector under the control of the inducible promoter nmt1, was
transformed into the fission yeast Schizosaccharomyces pombe. Transgenic
yeast cells that showed enhanced salt tolerance were selected. A total
of 69 such clones were identified from 10(6) of yeast transformants by
one-time screening. Functional assignment indicates that a large
percentage (81%) of these cDNAs encode for homologues of proteins that
are known to have roles in plant salt tolerance, while the other portion
(19%) codes for unknown proteins. Among the proteins encoded by the
known functional genes, the regulatory proteins account for a relatively
higher proportion. Northern blot analysis showed that the expression of
some isolated genes was salt stress inducible, whereas that of the
others was constitutive. These results indicate that fission yeast is a
simple and efficient system for functional and large-scale
identification of salt tolerance-related genes from other organisms.

Document Number: 270

During the past four decades, research efforts directed toward reducing
shell egg cholesterol content have centred on genetic selection or
alteration of the laying hens' diet with various nutrients, natural
products, non-nutritive factors, or pharmacological agents. However, the
vast majority of these experimental approaches elicited only minimal
changes (< 10%) at best or, as in the case of dietary azasterols and
triparanol, resulted in the unacceptable replacement of yolk cholesterol
by desmosterol. In contrast, when 3-hydroxy-3-methylglutaryl-coenzyme A
reductase inhibitors ('statins'), garlic paste, or pharmacological
amounts of copper were orally administered to chickens, yolk cholesterol
levels were reduced by up to 46%, 32%, or 34%, respectively. The
previous paper in this two-part series provided a justification for
low-cholesterol egg production and reviewed cholesterol metabolism in
the laying hen, avian embryonic cholesterol needs, and genetic and
nutritional approaches to chicken egg cholesterol reduction. The present
paper provides a comprehensive overview of the use of non-nutritive
dietary factors and select pharmacological compounds as egg
cholesterol-lowering agents, and discusses emerging strategies for
lowering the cholesterol content of shell eggs. With the recent advances
in avian genomics and transgenesis, it is anticipated that the greatest
advances in the area of yolk cholesterol reduction will be attained
through manipulation of key genes whose protein products mediate
intestinal sterol absorption, hepatic cholesterol and lipoprotein
synthesis, and/or lipoprotein uptake by growing oocytes. However, the
future commercial availability of low-cholesterol eggs produced by
transgenic hens ultimately will be influenced by the nature of the
regulatory environment for bioengineered foods, public acceptance, and
economics.

Document Number: 271

Transgenic expression of vitamin biosynthetic genes has been
investigated for over-production of these dietary supplements in
microorganisms and plants. In plants, successful efforts have been
reported with Vitamins A, C, E and B-9, however information is lacking
for other vitamins. Vitamin B-6 is an essential cofactor for numerous
enzymatic reactions, and has also been shown to be a potent antioxidant
involved in protecting phytopathogenic Cercospora fungi from their own
toxin, cercosporin. In this report, we transformed and expressed two
Vitamin B-6 biosynthetic genes (PDX1 and PDX2) isolated from Cercospora
nicotianae in cercosporin-sensitive organisms, tobacco and the fungal
species Aspergillus flavus and Neuraspora crassa. Our goal was to
determine if Vitamin B-6 levels could be increased by constitutive
expression of these genes, and if over-production confers resistance to
oxidative stresses induced by cercosporin and salinity stress. Elevated
Vitamin B-6 levels were observed in one tobacco line. For other lines
evaluated in this work, expression of PDX1 and PDX2 in transgenic
organisms did not result in a significant increase in Vitamin B-6
content over controls. Analysis of gene expression in tobacco indicated
that the lack of elevated B-6 content was not due to lack of enzymatic
activity, but to down-regulation of the endogenous tobacco genes
compounded with limited transgene expression. The single line with
elevated B-6 levels had higher expression of both the PDX1 and PDX2
transgenes compared to the other lines, and the observed increase on
Vitamin B-6 was correlated with higher enzyme activity. Consistent with
our inability to elevate cellular B-6 levels, only small changes were
observed in the response to either cercosporin or high salt, and most
transgenic individuals were as susceptible as controls. Compared to
tobacco lines transformed to express either PDX1 or PDX2 alone, half of
the transgenic tobacco lines expressing both genes were impaired in seed
germination and initial growth. However no correlation was observed
between the observed phenotype and Vitamin B-6 levels in seeds. This is
the first report on genetic engineering to manipulate the Vitamin B-6
pathway in plants. Our results suggest that genetic manipulation of the
Vitamin B-6 biosynthetic pathway is possible but is limited by
regulation of endogenous genes. (c) 2006 Elsevier Ireland Ltd. All
rights reserved.

Document Number: 272

The occurrence of intermixing, especially that resulting from
genetically modified (GM) species, is increasingly becoming a problem in
the delicate chain of feed and food quality control. Thus, a strategy is
needed for precisely quantifying the presence of intermixing. An
analytical assay based on real-time PCR has been developed; it can
ascertain the extent of unexpected intermixing of GM soybean with maize
meal. Three soybean-maize mix levels, with soybean intermix percentages
of, respectively, 0.1, 0.5, and 1%, were prepared to simulate samples
containing traces of soybean. As calibrator standards, ad hoc
multiple-target pGEM-T plasmids containing soybean and maize reference
genes in a 1:1 ratio were constructed. Four different maize endogenous
genes, alcohol dehydrogenase 1 (adh1), high-mobility group protein a
(hmga), invertase 1 (ivr1), and zein (zein), were assessed, each
combined with the soybean endogenous lectin 1 (lect1) gene. Plasmids
containing adh1-lect1 and zein-lect1 genes were found to be the most
reliable calibration systems for this analysis, providing precise and
accurate quantification results. Measuring the percentage of GM soybean
intermixing makes it possible to calculate the actual transgenic
component of the total sample.

Document Number: 273

Biofortification, the focus of the HarvestPlus program of the
Consultative Group on International Agriculture Research (CGIAR),
represents a potentially powerful tool to increase dietary intake of
essential nutrients in staple foods. This paper evaluates the
compatibility of biofortification with the preferred option of dietary
diversification and its potential impacts on the agricultural
biodiversity essential for long term sustainability. In poor countries,
biofortification requires increasing public investment in agricultural
research and infrastructure for success. Rather than cereal commodities,
biofortification for developing countries should focus on vegetatively
propagated species or in improving quality of coarse cereals, as well as
fodders. Community participatory approaches that identify local food
resources with nutritional, agronomic and economic advantages to
small-scale farmers could complement and set targets for
biofortification as one of many approaches to alleviate nutritional
deficiencies. Furthermore using agricultural biodiversity to reinforce
dietary diversity can help situate biofortification within the larger
context of sustainable food-based approaches. In this light, this paper
evaluates specific biofortification interventions from environmental,
sociocultural, political, economic, ethical, and biomedical
perspectives. (C) 2006 Elsevier Ltd. 

Document Number: 274

Since 1997, 18 studies with feeds from genetically modified plants (GMP)
in the nutrition of dairy cows, growing bulls, growing and finishing
pigs, laying hens, chicken for finishing as well as growing and laying
quails were conducted at the Federal Agricultural Research Centre (FAL)
in Braunschweig (Germany).
The majority of the experiments (16) were undertaken with GMP of the
so-called first generation (plants with input traits and without
substantial changes in composition) such as Bt-maize, Pat-maize,
Pat-sugar beet, Gt-soybean, Gt-potatoes and Bt-potatoes. Two studies
were carried out with GMP of the second generation (plants with output
traits or with substantial changes in their chemical composition) such
as an altered fatty acids profile in rapeseed or inulin potatoes. In all
experiments, feeds from GMP were compared with their isogenic
counterparts.
The iso- and transgenic feeds were analysed for their composition
(proximates, fibre fraction, amino acids, fatty acid pattern, minerals)
and undesirable substances (e.g., mycotoxins). Animal studies were
carried out for nutritional and safety assessment such as digestibility,
feed intake, health and performance of target animal species and quality
of food of animal origin. Reproduction was studied in a 10-generation
experiment with quails and a 4-generation experiment with laying hens.
Duration of experiments and number of animals were limited in some cases
due to small amounts of GM-feed available for experimentation.
Attention was drawn to the fate of DNA during feed processing (silage
making, oil extraction), in the digestive tract of animals (slaughtering
of animals 0, 4, 8, 12 and 24 It after feeding) and in the animal body
(samples from several organs and tissues).
In agreement with more than 100 animal studies available to date,
results show no significant differences in the nutritional value of
feeds from GMP of the first generation in comparison with non-GMP
varieties. To date, no fragments of recombinant DNA have been found in
any organ or tissue sample from animals fed GMP The lower content of
mycotoxins in Bt-maize and side effects in GMP of the second generation
are of safety concern.
The results indicate that routine feeding studies with target animal
species add little to nutritional assessment of feed from GMP of the
first generation, but they are of public interest and important for
safety assessment. These studies will play a more important role in
nutritional and safety assessment of feeds from GMP with output traits.
Proposals for such studies were made on the basis of previous
experiments. (c) 2006 Elsevier B.V. 

Document Number: 275

Since the commercialization of the first genetically modified (GM) crop
in 1996, the amount of arable land dedicated to the production of GM
feed has increased significantly. Despite widespread adoption of GM
foods and feeds, public perception of their safety remains mixed. To
provide consumers the opportunity for choice, some countries have
adopted mandatory labeling of GM foods and feeds when their adventitious
presence exceeds a defined threshold percentage. Methods for detecting
and quantifying GM plants in feeds include protein- and DNA-based
assays, but their sensitivity may be influenced by the techniques used
in feed processing. Interest in the consumption of transgenic protein
and DNA has prompted investigations of their fate within the
gastrointestinal tract of livestock and the potential to which
transgenes or their products may be incorporated into tissues.
Transgenic protein has not been detected in any animal tissues or
products. Fragments of DNA from endogenous, high-copy number chloroplast
genes from plants have been detected in poultry, pig and ruminant
tissues. Low-copy endogenous and transgenic DNA in animal tissues have
been detected but to a lesser extent than high-copy genes. Current
research suggests that the passage of dietary DNA fragments across the
intestinal wall is a natural physiological event, the likelihood of
which is dependent on their concentration in the feed. To date, the
transgenic traits approved for expression in crops used as feeds have
not posed a safety concern for livestock. (c) 2006 Elsevier B.V. All
rights reserved.

Document Number: 276

Although capture fisheries have experienced slow to stagnant growth in
recent years, the world population has been increasing, with subsequent
rises in demands for marine-based foods. One possibility for alleviating
potential food shortages and price increases is through aquaculture,
which has experienced rapid worldwide expansion. A major focus of
research in the aquaculture industry is on the use of biotechnology to
increase food availability and reduce production costs, specifically
through the manipulation of the genes and chromosomes of cultivated
species. Examples include transgenic fish with properties such as
increased growth rates, feed conversion efficiency, disease resistance,
cold tolerance, and improved metabolism of land-based plants. However,
use of transgenic organisms in aquaculture is a very controversial topic
due to a number of environmental and human health concerns such as
escapement and introduction of genetically modified organisms into the
food chain. In response, some transgenic research has also been focused
on inducing sterility to reduce the risk of transgenic organisms
breeding with wild species. A method of chromosome manipulation,
referred to as polyploidy, provides the option of creating sterile
organisms, some of which also exhibit increased growth rates. This
review paper will discuss recent advances in biotechnology research,
specifically in regards to the manipulation of genes and chromosomes,
for enhanced cultivation of fish and invertebrates. Major environmental
and human health concerns will also be addressed.

Document Number: 277

A new transgenic cotton producing the Vegetative Insecticidal Protein
(Vip) is being developed to control Lepidopteran pests, especially
Helicoverpa larvae. Before its introduction its efficacy against
Helicoverpa larvae under field conditions needs to be confirmed, and any
non-target effects it may have on the arthropod community need to be
identified. We conducted field trials to compare the arthropod community
in unsprayed conventional (Sicala 40) and Vip (Coker 312 Vip3A, event
102) cotton using visual searches, beatsheets, and suction samplers at 2
sites in Australia. At both sites, Vip controlled Helicoverpa larvae
leading to much higher boll counts. There were no major differences in
either species richness or diversity of the beneficial and non-target
communities between Vip and conventional cotton, although cotton
cultivar accounted for 2-7% of the variance in arthropod communities.
There was no detrimental effect of Vip cotton on egg parasitoids. The
number of predatory beetles and the pest mirid Creotiades dilutus(Stal)
was higher in the Vip, although the increase in mirids was probably the
result of more food (bolls) in the Vip crop. In a small plot experiment,
we found higher numbers of whitefly in Vip, but this may be driven by
differences in leaf hair between the cotton cultivars. Vip cotton
appeared to have little effect on the arthropod community other than on
Helicoverpa. As such it has the potential to be a useful tool in the
management of Helicoverpa and may relieve resistance pressure on
existing Bt cultivars (transgenic cotton containing genes for
insecticidal Cry proteins), thereby increasing the durability of both
technologies.

Document Number: 278

A new transgenic cotton producing the Vegetative Insecticidal Protein
(Vip) is being developed to control Lepidopteran pests, especially
Helicoverpa larvae. Before its introduction its efficacy against
Helicoverpa larvae under field conditions needs to be confirmed, and any
non-target effects it may have on the arthropod community need to be
identified. We conducted field trials to compare the arthropod community
in unsprayed conventional (Sicala 40) and Vip (Coker 312 Vip3A, event
102) cotton using visual searches, beatsheets, and suction samplers at 2
sites in Australia. At both sites, Vip controlled Helicoverpa larvae
leading to much higher boll counts. There were no major differences in
either species richness or diversity of the beneficial and non-target
communities between Vip and conventional cotton, although cotton
cultivar accounted for 2-7% of the variance in arthropod communities.
There was no detrimental effect of Vip cotton on egg parasitoids. The
number of predatory beetles and the pest mirid Creotiades dilutus (Stal)
was higher in the Vip, although the increase in mirids was probably the
result of more food (bolls) in the Vip crop. In a small plot experiment,
we found higher numbers of whitefly in Vip, but this may be driven by
differences in leaf hair between the cotton cultivars. Vip cotton
appeared to have little effect on the arthropod community other than on
Helicoverpa. As such it has the potential to be a useful tool in the
management of Helicoverpa and may relieve resistance pressure on
existing Bt cultivars (transgenic cotton containing genes for
insecticidal Cry proteins), thereby increasing the durability of both
technologies.

Document Number: 279

Protein expression in a maize hybrid flour (hereafter called WT) and its
corresponding transgenic version resistant to European corn borer
(hereafter called BT and carrying a gene encoding for the Bacillus
thuringiensis insecticidal protein Cry1Ab) has been studied by means of
two-dimensional gel electrophoresis coupled with mass spectrometry. This
comparison has been chosen as a model to verify proteomics capability in
detecting unexpected differences between near-isogenic lines (differing
in the ideal case only for the presence of the transgenic protein). Some
unpredictable differences were detected: i) glucose and ribitol
dehydrogenase spot was unique of BT maize; ii) endochitinase A spot was
unique of WT maize; iii) triosephosphate isomerase 1 and one spot of
globulin-1 S were overexpressed while cytosolic 3-phosphoglycerate
kinase and one spot of aldose reductase were down-regulated in BT maize
with respect to WT. These results outline the potential of the new
non-targeted "-omics" technologies (in particular proteomics) in the
detection of unexpected, unintended and unwanted variations in
Genetically Modified (GM) versus non-GM food comparison and suggest the
possible employment of these technologies in substantial equivalence
evaluation. Moreover, a reference map for maize flour was built: forty
spots, corresponding to twenty-five different proteins, were
successfully identified.

Document Number: 280

According to the information reported by the WHO, the genetically
modified (GM) products that are currently on the international market
have all passed risk assessments conducted by national authorities.
These assessments have not indicated any risk to human health. In spite
of this clear statement, it is quite amazing to note that the review
articles published in international scientific journals during the
current decade did not find, or the number was particularly small,
references concerning human and animal toxicological/health risks
studies on GM foods. In this paper, the scientific information
concerning the potential toxicity of GM/transgenic plants using the
Medline database is reviewed. Studies about the safety of the potential
use of potatoes, corn, soybeans, rice, cucumber, tomatoes, sweet pepper,
peas, and canola plants for food and feed were included. The number of
references was surprisingly limited. Moreover, most published studies
were not performed by the biotechnology companies that produce these
products. This review can be concluded raising the following question:
where is the scientific evidence showing that GM plants/food are
toxicologically safe?.

Document Number: 281

Potential safety issues have been raised with the development and
release of virus-resistant transgenic plants. This review focuses on
safety assessment with a special emphasis on crops that have been
commercialized or extensively tested in the field such as squash,
papaya, plum, grope, and sugar beet. We discuss topics commonly
perceived to be of concern to the environment and to human
health-heteroencapsidation, recombination, synergism, gene flow, impact
on nontarget organisms, and food safety in terms of allergenicity. The
wealth of field observations and experimental data is critically
evaluated to draw inferences on the most relevant issues. We also
express inside views on the safety and benefits of virus-resistant
transgenic plants, and recommend realistic risk assessment approaches to
assist their timely deregulation and release.

Document Number: 282

Genetic modification (GM) of plants has great potential in the
production of food and industrial compounds, and in molecular pharming.
One of the greatest public concerns regarding this technology is
effective pollen flow, in which wind- or insect-borne transgenic pollen
is able to fertilise either non-GM crops of the same species, or closely
related weed species, and lead to viable seed formation. In this paper
we describe a novel concept, based on epigenetic inheritance
(imprinting) and post-transcriptional gene silencing (PTGS)/RNA
interference (RNAi), designed to prevent transgene escape via pollen
flow from transgenic plants. A key advantage of this strategy is that it
would allow all seeds from self-pollinated transgenic plants to be
harvested and re-sown, without the need for specific treatments, while
retaining all of the transgenes present in the parent. Thus, this
strategy is not a Genetic Use Restriction Technology (GURT) and if
implemented would not prevent seed saving by end-users.

Document Number: 283

Rice is the most amenable crop plant for genetic manipulation amongst
monocots due to its small genome size, enriched genetic map,
availability of entire genome sequence, and relative ease of
transformation. Improvement in agronomic traits of rice is bound to
affect a sizeable population since it is a primary source of sustenance.
Recent advances like use of 'clean gene' technology or matrix attachment
regions would help improve rice transformation. Function of several
novel genes and their promoters has been analyzed in transgenic rice.
Significant progress has been made in introducing traits like herbicide,
biotic stress and abiotic stress tolerance. Attempts also have been made
to enhance nutritional characteristics of the grain and yield.
Identification of genes controlling growth and development can be used
to modify plant architecture and heading period. Transgenic rice can
serve as a biofactory for the production of molecules of pharmaceutical
and industrial utility. The drive to apply transgenic rice for public
good as well as commercial gains has fueled research to an all time
high. Successful field trials and biosafety of transgenic rice have been
reported. This would act as a catalyst for greater acceptance of
genetically modified food crops. The lessons learnt from rice can be
extended to other cereals thereby opening new opportunities and
possibilities.

Document Number: 284

Proanthocyanidins (PAs) are oligomeric plant natural products mostly
derived from epicatechin and/or catechin monomers. In studies aimed at
engineering PAs into plant tissues that do not naturally make these
compounds, we have expressed PA biosynthetic and regulatory genes in
tobacco, alfalfa (Medicago sativa) and the model legume Medicago
truncatula. Because engineered tannins may be produced in small
quantities and it is often necessary to screen many independent plant
lines, we have developed an improved, highly sensitive method to
quantify and determine the composition of oligomeric PAs in plant
extracts. The method involves normal-phase HPLC separation of
semi-purified PAs followed by post-column reaction with the PA-specific
reagent DMACA (dimethylaminocinnamaldehyde). This procedure allows for
accurate and sensitive quantification of individual oligomeric PAs and,
unlike currently used methods, does not require exhaustive sample
preparation and clean-up. Compositional data are shown for genetically
engineered PAs in tobacco and alfalfa.

Document Number: 285

Concerns over the safety of GM crops have led to detailed comparisons of
their composition and performance with those of conventionally bred
crops, under both field and glasshouse conditions. Such studies of wheat
have shown that it is possible to develop GM lines which are
substantially equivalent to conventional varieties. This information is
important to inform the debate on the future development of GM food
crops.

Document Number: 286

Transgenic rice seeds, which express a hybrid peptide comprising seven
predominant human T cell epitopes (7Crp) derived from Japanese cedar
pollen allergens, have been shown to function as an effective edible
vaccine for the control of pollen allergen-induced responses. In this
study, we characterized biochemical properties of transgenic seeds
expressing the 7Crp peptide. The levels of chemical compositions, such
as carbohydrate, protein, lipid, amino acid, fatty acid, mineral, and
vitamin, were substantially equivalent between transgenic 7Crp and its
nontransgenic counterpart seeds. The contents of three major allergenic
proteins in transgenic seeds were not enhanced by expression of the 7Crp
peptide when compared with those of nontransgenic seeds. The 7Crp
peptide expressed in seeds was susceptible to simulated
gastric/intestinal fluids. N-Glycosylation was not observed in the 7Crp
peptide sequence. These results indicate that transgenic 7Crp seeds are
substantially equivalent to nontransgenic parental seeds except for the
presence of the 7Crp peptide.

Document Number: 287

Recent reports, workshops and meetings on GM (Genetically Modified)
crops tend to characterize GM food production as a solution to Africa's
food crisis. However, GM crops are currently grown commercially in only
one country in Africa - South Africa. Biotechnology tools range from
tissue culture to molecular breeding and genetic engineering. This paper
focuses on GM crop improvement and analyzes the development of seven GM
crops (six food staples and cotton) over the past 15 years in Africa.
The case studies reveal a number of unexpected scientific, legal,
economic and political barriers to the development of GM crops and long
delays in developing and implementing national biosafety regulations and
guidelines. We conclude that most GM crops are at least 1015 years or
longer from reaching smallholder farmers in Africa. During this time
special attention should be given to strengthening conventional plant
breeding programs in NARS (National Agricultural Research Systems),
African universities and the CGIAR. Biotechnology approaches must be
nested and integrated into plant breeding programs. Special attention
should also be given to raising public awareness of biotechnology,
mobilizing political support and commitment to strengthening African
capacity in biotechnology, biosafety, food safety and IPR (Intellectual
Property Rights) and mounting long-term training programs to train the
next generation of African plant breeders and GM crop specialists. (c)
2006 Elsevier Ltd. 

Document Number: 288

The stability of the inserted transgenes and particularly the junction
regions of transgenic events is a concern of food labeling, traceability
and post release monitoring, as these regions are used for development
of event-specific DNA-based detection methods. During the standard
agricultural breeding practices, the transgenic lines can be exposed to
completely different conditions than those in the laboratory
environment. Some of these conditions have the potential to affect the
stability of the transgenic locus and the surrounding DNA. As tissue
culture is recognized as a stressful and mutagenic factor, we have
analyzed the effect of this process on the stability of the junction
regions at nucleotide level in five Arabidopsis thaliana transgenic
lines in comparison with the respective integration loci in ColO and C24
ecotypes. No indication of any kind of alteration at nucleotide level of
the junctions was found. The relevance of the stability of the
plant-T-DNA junction regions for application of the DNA-based methods in
commercial transgenic plants is discussed.

Document Number: 289

Genetically modified rice varieties developed in China are close to
approval for agricultural cultivation and production. However, so far no
method has been reported for specific detection of transgenic varieties
of this crop. In the present study, rice seeds assumed to consist of
field-tested Bt rice ('Anti-pest Shanyou 63 degrees and Anti-pest Jinyou
63 degrees) were used as reference material to determine transgenic DNA
sequences. The transition between the cryIA(b) and cryIA(c) fusion gene
and the nopaline synthase terminator (nos) sequence was used to develop
a construct-specific real-time PCR based detection method. This Bt rice
specific detection system was combined with a recently published
quantitative real-time PCR method for the rice-specific (Oryza sativa
L.) reference gene gos9. The complete PCR assay for detection of
transgenic Bt rice was in-house validated and the limit of
quantification was found to be below 0.1% Bt rice relative to the rice
content. Application of the PCR assay should allow more precise
detection of transgenic rice varieties in imported food products which
are so far not approved in the EU.

Document Number: 290

In this study, the comparative effects of transgenic corn (Mon 810 and
Event 176) and isogenic corn (DK729) were investigated for their
influence on in vitro rumen fermentation. This study consisted of three
treatments with 0.25 g rice straw, 0.25 g of corn (Mon810/Event176/DK
729) mixed with 30 ml rumen fluid-basal medium in a serum bottle. They
were prepared in oxygen free conditions and incubated at 39 degrees C in
a shaking incubator. The influence of transgenic corn on the number of
bacterial population, E succinogenes (cellulolytic) and S. bovis
(amylolytic), was quantified using RT-PCR. Fermentative parameters were
measured at 0, 2, 4, 8, 12 and 24 h and substrate digestibility was
measured at 12 and 24 h. No significant differences were observed in
digestibility of dry matter, NDF, ADF at 12 and 24 h for both transgenic
and isogenic form of corns (p > 0.05) as well as in fermentative
parameters. Fluid pH remained unaffected by hybrid trait and decreased
with VFA accumulation as incubation time progressed. No influence of
corn trait itself was seen on concentration of total VFA, acetic,
propionic, butyric and valeric acids. There were no significant
differences (p < 0.05) in total gas production, composition of gas
(methane and hydrogen) at all times of sampling, as well as in NH3-N
production. Bacterial quantification using RT-PCR showed that the
population number was not affected by transgenic corn. From this study
it is concluded that transgenic corn (Mon810 and Event 176) had no
adverse effects on rumen fermentation and digestibility compared to
isogenic corn. However, regular monitoring of these transgenic feeds is
needed by present day researchers to enable consumers with the option to
select their preferred food source for animal or human consumption.

Document Number: 291

Plants have emerged as a modem production system to produce recombinant
proteins-antigens that can be used as subunit vaccines. The ideal plant
candidate for this purpose should be capable to sustain high levels of
expression of foreign proteins without adverse effects on its growth and
development. It is also essential that it has large biomass, is edible
and suitable for long-term storage and delivery.
This work is a part of an effort to develop Cruciferae-based production
system using transgenic vegetable plants collard and cauliflower.
Several parameters were tested and optimized to achieve an efficient
stable transformation of these recalcitrant species with constructs
containing expression cassettes for the known viral antigens. Using the
original procedure we obtained transgenic collard cv Morris Heading that
express high levels of smallpox vaccine candidate (B5) in leaves and
retain its normal phenotype. Transgenic cauliflower plants cv Early
Snowball were obtained in similar procedure and have shown detectable
amounts of SARS coronavirus spike-protein (SARS-CoV S1) in floret tissue
of mature curd.
To our knowledge, this is the first report on generation of transgenic
collard plants ever and the first successful attempt to use these
vegetables for production of pharmaceutical proteins. (c) 2006 Elsevier
Ireland Ltd. 

Document Number: 292

The effect of the digestion process in the gastro-intestinal tract (GIT)
of animal models on the fate and integrity of plant DNA has been widely
evaluated since DNA availability and integrity is a key factor for
hypothetical horizontal gene transfer of recombinant DNA from GM
crop-derived feeds to animal and human gut microflora. In this study,
plant DNA sequences from high and low copy number genes were monitored
in GIT and tissues of buffaloes and rabbits. Using a real-time PCR
approach to track plant DNA in animal samples, we demonstrated the
persistence of fragmented plant DNA blood and tissues of buffaloes and
rabbits raised with conventional feeding. (c) 2006 Elsevier B.V. All
rights reserved.

Document Number: 293

Potatoes, members of the Solanaceae plant family, serve as major,
inexpensive low-fat food sources providing energy (starch), high-quality
protein, fiber, and vitamins. Potatoes also produce biologically active
secondary metabolites, which may have both adverse and beneficial
effects in the diet. These include glycoalkaloids, calystegine
alkaloids, protease inhibitors, lectins, phenolic compounds, and
chlorophyll. Because glycoalkaloids are reported to be involved in
host-plant resistance and to have a variety of adverse as well as
beneficial effects in cells, animals, and humans, a need exists to
develop a clearer understanding of their roles both in the plant and in
the diet. To contribute to this effort, this integrated review presents
data on the (a) history of glycoalkaloids; (b) glycoalkaloid content in
different parts of the potato plant, in processed potato products, and
in wild, transgenic, and organic potatoes; (c) biosynthesis,
inheritance, plant molecular biology, and glycoalkaloid-plant
phytopathogen relationships; (d) dietary significance with special focus
on the chemistry, analysis, and nutritional quality of low-glycoalkaloid
potato protein; (e) pharmacology and toxicology of the potato
glycoalkaloids comprising alpha-chaconine and alpha-solanine and their
hydrolysis products (metabolites); (f) anticarcinogenic and other
beneficial effects; and (g) possible dietary consequences of concurrent
consumption of glycoalkaloids and other biologically active compounds
present in fresh and processed potatoes. An enhanced understanding of
the multiple and overlapping aspects of glycoalkaloids in the plant and
in the diet will benefit producers and consumers of potatoes.

Document Number: 294

This review highlights recombinant DNA technology as a powerful tool to
enhance the gene pools of sorghum and pearl millet crops regarded as
jewels of Africa. Although important advances in the improvement of
these species have been made by classical breeding and modern marker
assisted selection, genetic manipulation and in vitro culture allows the
gene pool to be broadened beyond that normally available for improvement
by allowing the transfer of genes which control well-defined traits
between species. The current state of sorghum and millet transformation
technology is summarised and applications in the improvement of
nutritional quality and the resistance to pathogens and pests for crops
grown in Africa and Asia is discussed. Regulatory aspects including gene
flow and future prospects are also discussed. (c) 2006 Elsevier Ltd. All
rights reserved.

Document Number: 295

Lactoferrin is an iron-binding glycoprotein present in milk as well as
other exocrine secretions and neutrophil granules in mammals.
Lactoferrin is considered to be an important host defense molecule and
has a diverse range of physiological functions such as
antimicrobial/antiviral activities, immunomodulatory activity, and
antioxidant activity. During the past decade, it has become evident that
oral administration of lactoferrin exerts several beneficial effects on
the health of humans and animals, including anti-infective, anticancer,
and anti-inflammatory effects. This has enlarged the application
potential of lactoferrin as a food additive. The technology of producing
bovine lactoferrin on a factory scale was established over 20 years ago.
Bovine lactoferrin is purified by cation-exchange chromatography from
bovine skim milk or whey, and is commercially available from several
suppliers. Recombinant human lactoferrin is produced by Aspergillus
niger, transgenic cows, and rice, and its efficacy is being evaluated.
In this article, we review basic research and technological aspects of
the application of lactoferrin. (c) 2006 Elsevier Ltd. All rights
reserved.

Document Number: 296

Compositional information for lignins in food is rare and concentrated
on cereal grains and brans. As lignins are suspected to have important
health roles in the dietary fiber complex, the confusing current
information derived from nonspecific lignin determination methods needs
to be augmented by diagnostic structural studies. For this study, lignin
fractions were isolated from kiwi, pear, rhubarb, and, for comparison,
wheat bran insoluble dietary fiber. Clean pear and kiwi lignin isolates
allowed for substantive structural profiling, but it is suggested that
the significance of lignin in wheat has been overestimated by reliance
on nonspecific analytical methods. Volume integration of NMR contours in
two-dimensional C-13-H-1 correlation spectra shows that pear and wheat
lignins have comparable guaiacyl and syringyl contributions and that
kiwi lignins are particularly guaiacyl-rich (similar to 94% guaiacyl)
and suggest that rhubarb lignins, which could not be isolated from
contaminating materials, are as syringyl-rich (similar to 96% syringyl)
as lignins from any known natural or transgenic fiber source. Typical
lignin structures, including those newly NMR-validated (glycerols,
spirodienones, and dibenzo-dioxocins), and resinols implicated as
possible mammalian lignan precursors in the gut are demonstrated via
their NMR correlation spectra in the fruit and vegetable samples. A
novel putative benzodioxane structure appears to be associated with the
kiwi lignin. It is concluded that the fruits and vegetables examined
contain authentic lignins and that the detailed structural analysis
exposes limitations of currently accepted analytical methods.

Document Number: 297

The complete sequence of the genomes of two model plants is now
available together with technology for. the parallel analysis of
thousands of genes, the proteins that they encode and the metabolites
whose production they control. This review focuses on application of
these technologies to engineer output characteristics of crops
specifically relevant to non-food use. Following a brief introduction to
the resources available for genomic analysis, some examples are provided
of how genomics is enabling both transgenic and non-GM manipulation of
agricultural materials such as carbohydrates, oils, fibres and
speciality chemicals for use as industrial feedstocks. (c) 2006 Society
of Chemical Industry.

Document Number: 298

Genetically modified plants must be approved before release in the
European Union, and the approval is generally based upon a comparison of
various characteristics between the transgenic plant and a conventional
counterpart. As a case study, focusing on safety assessment of
genetically modified plants, we here report the development and
characterisation of six independently transformed Arabidopsis thaliana
lines modified in the flavonoid biosynthesis. Analyses of integration
events and comparative analysis for characterisation of the intended
effects were performed by PCR, quantitative Real-time PCR, and High
Performance Liquid Chromatography. Analysis by cDNA microarray was used
as a non-targeted approach for the identification of potential
unintended effects caused by the transformation. The results revealed
that, although the transgenic lines possessed different types of
integration events, no unintended effects were identified. However, we
found that the majority of genes showing differential expression were
identified as stress-related genes and that environmental conditions had
a large impact on the expression of several genes, proteins, and
metabolites. We suggest that the microarray approach has the potential
to become a useful tool for screening of unintended effects, but state
that it is crucial to have substantial information on the natural
variation in traditional crops in order to be able to interpret "ornics"
data correctly within the framework of food safety assessment strategies
of novel plant varieties, including genetically modified plant
varieties.

Document Number: 299

Resistance to antibiotics mediated by selectable marker genes remains a
powerful selection tool for transgenic event production. However,
regulatory agencies and consumer concerns favor these to be eliminated
from food crops. Several excision systems exist but none have been
optimized or shown to be functional for clonally propagated crops. The
excision of the nptII gene conferring resistance to kanamycin has been
achieved here using a gene construct based on a heat-inducible cre gene
producing a recombinase that eliminates cre and nptII genes flanked by
two loxP sites. First-generation regenerants with the Cre-loxP system
were obtained by selection on kanamycin media. Following a heat
treatment, second generation regenerants were screened for excision by
PCR using nptII, cre, and T-DNA borders primers. Excision efficiency
appeared to be at 4.7% depending on the heat treatment. The footprint of
the excision was shown by sequencing between T-DNA borders to correspond
to a perfect recombination event. Selectable marker-free sprouts were
also obtained from tubers of transgenic events when submitted to similar
heat treatment at 4% frequency. Spontaneous excision was not observed
out of 196 regenerants from untreated transgenic explants. Biosafety
concerns are minimized because the expression of cre gene driven by the
hsp70 promoter of Drosophila melanogaster was remarkably low even under
heat activation and no functional loxP site were found in published
Solanum sequence database. A new plant transformation vector pCIP54/55
was developed including a multiple cloning site and the self-excision
system which should be a useful tool not only for marker genes in potato
but for any gene or sequence removal in any plant.

Document Number: 300

The main objective of this study was to introduce the artificial neural
network (ANN) technique into the field of food analysis. The specific
purpose was to evaluate the lipid distribution of Bt-176 transgenic
maize compared to that of conventional maize. The crude oil extracted
from the grains of genetically modified maize (Bt-176) and nontransgenic
maize was characterized in terms of the fatty acid, sterol, tocopherol
distribution as well as the lipid classes and unsaponifiable level. The
content of total lipids was within the range of 3.21-3.40% of grain dry
matter. Fractionation of lipids into polar and nonpolar classes showed
that the transgenic maize (Bt-176) contained more polar lipids than the
control maize. In general, results obtained from lipid distribution
analysis showed that, except for a few minor differences, the grains of
Bt-176 maize were comparable in composition to that of the control
maize. On the other hand, the analytical data have been elaborated by
supervised pattern recognition technique (ANN) in order to classify
genetically modified maize (Bt-176) and conventional maize as well as to
authenticate the origin of the samples. (c) 2006 Elsevier Inc. All
rights reserved.

Document Number: 301

Gene flow via outcrossing from transgenic plants to relatives will be
one of the most important concerns to grow of the transgenic chickpea
(Cicer arietinum L.) in European Union (EU). This report is therefore
focused on spontaneous outcrossing rate in chickpea. A total of 39
kabuli type mutants with white flower and one desi type with pink flower
were grown to estimate spontaneous outcrossing rate. Outcrossing rate
ranged from 0.0 to 1.25% in mutant materials. Since labelling threshold
for transgenic contamination in food and feed in European Union (EU) is
0.9%, outcrossing rate of 1.25% is higher than threshold of 0.9% in EU,
and this result suggests that cultivation of transgenic chickpea will be
under high risk to be contaminated chickpeas in neighbourhood fields.

Document Number: 302

Recently, DNA-based techniques became very common for the detection of
genetically modified organisms (GMOs) in food products. For rapid and
easy detection of GMOs, polymerase chain reaction (PCR) screening
methods.. which amplify common transgenic elements, are applied in
routine analysis. Incorporation of PCR and membrane method introduced in
this study offer an alternative detection of GMOs. I it this study, a
total of 32 samples and three certified reference materials were tested
for the existence of the 35S promoter of cauliflower mosaic virus (CaMV)
and 5-enol-pyruvyl-shikimate-3-phosphate synthase (EPSPS) gene residues.
Dot blot screening system introduced in this study can be routinely used
as a semi-quantitative screening of GMOs. (c) 2005 Elsevier Ltd. All
rights reserved.

Document Number: 303

Flavonoids are a large family of plant polyphenolic secondary
metabolites. Although they are widespread throughout the plant kingdom,
some flavonoid classes are specific for only a few plant species. Due to
their presumed health benefits there is growing interest in the
development of food crops with tailor-made levels and composition of
flavonoids, designed to exert an optimal biological effect. In order to
explore the possibilities of flavonoid engineering in tomato fruits, we
have targeted this pathway towards classes of potentially healthy
flavonoids which are novel for tomato. Using structural flavonoid genes
(encoding stilbene synthase, chalcone synthase, chalcone reductase,
chalcone isomerase and flavone synthase) from different plant sources,
we were able to produce transgenic tomatoes accumulating new
phytochemicals. Biochemical analysis showed that the fruit peel
contained high levels of stilbenes (resveratrol and piceid),
deoxychalcones (butein and isoliquiritigenin), flavones
(luteolin-7-glucoside and luteolin aglycon) and flavonols (quercetin
glycosides and kaempferol glycosides). Using an online high-performance
liquid chromatography (HPLC) antioxidant detection system, we
demonstrated that, due to the presence of the novel flavonoids, the
transgenic tomato fruits displayed altered antioxidant profiles. In
addition, total antioxidant capacity of tomato fruit peel with high
levels of flavones and flavonols increased more than threefold. These
results on genetic engineering of flavonoids in tomato fruit demonstrate
the possibilities to change the levels and composition of health-related
polyphenols in a crop plant and provide more insight in the genetic and
biochemical regulation of the flavonoid pathway within this worldwide
important vegetable.

Document Number: 304

The present paper addresses the question of thermotolerance of in planta
synthesized heterologous enzymes using phytase as a model. Two
individual transgenic wheat materials expressing an Aspergillus
fumigatus phytase with a low denaturation temperature (62.5 degrees C)
but a high refolding capacity, and a rationally designed consensus
phytase engineered to a high denaturation temperature (89.3 degrees C),
were evaluated. High levels of endosperm specific expression were
ensured by the wheat high molecular weight glutenin 1DX5 promoter.
Immunodetection at the light and electron microscopical level shows
unequivocally that the heterologous phytase is deposited in the vacuole,
albeit that the transformation constructs were designed for secretion to
the apoplast. Evaluation of heat stability properties and kinetic
properties unraveled that, under these deposition conditions, heat
stability based on high unfolding temperature is superior to high
refolding capacity and represents a realistic strategy for improving
phosphate and mineral bioavailability in cereal-based feed and food.

Document Number: 305

Background: Potato is a major staple food, and modification of its
provitamin content is a possible means for alleviating nutritional
deficiencies. beta-carotene is the main dietary precursor of vitamin A.
Potato tubers contain low levels of carotenoids, composed mainly of the
xanthophylls lutein, antheraxanthin, violaxanthin, and of xanthophyll
esters. None of these carotenoids have provitamin A activity.
Results: We silenced the first dedicated step in the beta-epsilon-branch
of carotenoid biosynthesis, lycopene epsilon cyclase (LCY-e), by
introducing, via Agrobacterium-mediated transformation, an antisense
fragment of this gene under the control of the patatin promoter. Real
Time measurements confirmed the tuber-specific silencing of Lcy-e.
Antisense tubers showed significant increases in beta-beta-carotenoid
levels, with beta-carotene showing the maximum increase (up to 14-fold).
Total carotenoids increased up to 2.5-fold. These changes were not
accompanied by a decrease in lutein, suggesting that LCY-e is not
rate-limiting for lutein accumulation. Tuber-specific changes in
expression of several genes in the pathway were observed.
Conclusion: The data suggest that epsilon-cyclization of lycopene is a
key regulatory step in potato tuber carotenogenesis. Upon tuber-specific
silencing of the corresponding gene, beta-beta-carotenoid and total
carotenoid levels are increased, and expression of several other genes
in the pathway is modified.

Document Number: 306

Evidence for substantial environmental influences on health and food
safety comes from work with environmental health indicators which show
that agroenvironmental practices have direct and indirect effects on
human health, concluding that "the quality of the environment influences
the quality and safety of foods" [Fennema, O. Environ. Health Perspect.
1990, 86, 229- 232). In the field of genetically modified organisms
(GMOs), Codex principles have been established for the assessment of GM
food safety and the Cartagena Protocol on Biosafety outlines
international principles for an environmental assessment of living
modified organisms. Both concepts also contain starting points for an
assessment of health/ food safety effects of GMOs in cases when the
environment is involved in the chain of events that could lead to
hazards. The environment can act as a route of unintentional entry of
GMOs into the food supply, such as in the case of gene flow via pollen
or seeds from GM cropss, but the environment can also be involved in
changes of GMO-induced agricultural practices with relevance for health/
food safety. Examples for this include potential regional changes of
pesticide uses and reduction in pesticide poisonings resulting from the
use of Bt crops or influences on immune responses via cross-reactivity.
Clearly, modern methods of biotechnology in breeding are involved in the
reasons behind the rapid reduction of local varieties in agrodiversity,
which constitute an identified hazard for food safety and food security.
The health/ food safety assessment of GM foods in cases when the
environment is involved needs to be informed by data from environmental
assessment. Such data might be especially important for hazard
identification and exposure assessment. International organizations
working in these areas will very likely be needed to initiate and enable
cooperation between those institutions responsible for the different
assessments, as well as for exchange and analysis of information. An
integrated assessment might help to focus and save capacities in highly
technical areas such as molecular characterization or profiling, which
are often necessary for both assessments. In the area of establishing
international standards for traded foods, such as for the newly created
Standards in Trade and Development Facility (STDF), an integrated
assessment might help in the consideration of important environmental
aspects involved in health and food safety. Furthermore, an established
integrated view on GMOs may create greater consumer confidence in the
technology.

Document Number: 307

A review is presented demonstrating that biospecific interaction
analysis, using surface plasmon resonance (SPR) and biosensor
technologies is a simple, rapid, and automatable approach to detect
genetically modified organisms (GMOs). Using SPR, we were able to
monitor in real-time the hybridization between oligonucleotide or
polymerase chain reaction (PCR)-generated probes and target
single-stranded PCR products obtained by using as substrates DNA
isolated from normal or transgenic soybean and maize. This procedure
allows a one-step, nonradioactive detection of GMOs. PCR-generated
probes are far more efficient in detecting GMOs than are
oligodeoxy-ribonucleotide probes. This is expected to be a very
important parameter, because information on low percentage of GMOs is of
great value. Determination of the ability of SPR-based analysis to
quantify GMOs should be considered a major research field for future
studies, especially for the analyses of food supplies.

Document Number: 308

Biotechnology has permitted the modification of agricultural materials
in a very precise way to improve productivity and yields. Polymerase
chain reaction (PCR)-based methods have been the first choice of most
analytical laboratories for routine use in the detection of genetically
modified organisms (GMO) and their derived products. These methods rely
on the amplification of transgenic sequences and quantification of the
transgenic DNA by comparison with an amplified reference gene. This
paper describes the specific primers and probe for the cotton
stearoyl-ACP desaturase (sad1) gene, and PCR cycling conditions suitable
for the use of this sequence, which acts as an endogenous reference gene
in both qualitative and quantitative PCR assays. The two methods were
tested with 18 cotton varieties and identical amplification products
were obtained with all of them. No amplification products were detected
when DNA samples from other species, including soybean, rapeseed,
tobacco, maize, tomato, potato, cucumber, pea, red pepper, sunflower,
sesame, rice, peach, banana, apple, pumpkin, barley and carrot, were
used as templates, which demonstrates that this system is specific for
cotton. In real-time quantitative PCR analysis, the detection limit was
as low as 6 pg of DNA, which indicates that this method is suitable for
application to processed food samples that contain very low copies of
target DNA. Southern blot analysis confirmed that the sad1 gene was a
single copy in the tested cotton varieties. (c) 2006 Society of Chemical
Industry.

Document Number: 309

Plant biotechnology can make important contributions to food security
and nutritional improvement. For example, the development of 'Golden
Rice' by Professor Ingo Potrykus was a milestone in the application of
gene technology to deliver both increased nutritional qualities and
health improvement to wide sections of the human population. Mineral
nutrient and protein deficiency as well as food security remain the most
important challenges for developing countries. Current projects are
addressing these issues in two major staple crops, cassava (Manihot
esculenta Crantz) and rice. The tropical root crop cassava is a major
source of food for approximately 600 million of the population
worldwide. In sub-Saharan Africa > 200 million of the population rely on
cassava as their major source of dietary energy. The nutritional quality
of the cassava root is not sufficient to meet all dietary needs. Rice is
the staple food for half the world population, providing approximately
20% of the per capita energy and 13% of the protein for human
consumption worldwide. In many developing countries the dietary
contributions of rice are substantially greater (29 center dot 3%
dietary energy and 29 center dot 1% dietary protein). The current six
most popular 'mega' rice varieties (in terms of popularity and acreage),
including Chinese hybrid rice, have an incomplete amino acid profile and
contain limited amounts of essential micronutrients. Rice lines with
improved Fe contents have been developed using genes that have functions
in Fe absorption, translocation and accumulation in the plant, as well
as improved Fe bioavailability in the human intestine. Current
developments in biotechnology-assisted plant improvement are reviewed
and the potential of the technology in addressing human nutrition and
health are discussed.

Document Number: 310

A multiplex DNA microarray chip was developed for simultaneous
identification of nine genetically modified organisms (GMOs), five plant
species and three GMO screening elements, i.e. the 35S promoter, the nos
terminator and the nptII gene. The chips also include several controls,
such as that for the possible presence of CaMV. The on-chip detection
was performed directly with PCR amplified products. Particular emphasis
was placed on the reduction of the number of PCR reactions required and
on the number of primers present per amplification tube. The targets
were biotin labelled and the arrays were detected using a colorimetric
methodology. Specificity was provided by specific capture probes
designed for each GMO and for the common screening elements. The
sensitivity of the assay was tested by experiments carried out in five
different laboratories. The limit of detection was lower than 0.3% GMO
for all tests and in general around 0.1% for most GMOs. The chip
detection system complies with the requirements of current EU
regulations and other countries where thresholds are established for the
labelling of GMO.

Document Number: 311

To ensure food production appropriate for poulation requirements, in
terms of quantity. quality, and the health security, while respecting
the principles of sustainable development for the planet, crop
protection, which today remains principally agrochemical, must change or
it will aggravate an already disquieting economc and balance sheet. The
alternative proposed at the end of the 1960s - the concept of integrated
pest management - has been successful only in the most critical
situations: fruit orchards and greenhouses. The transgenic solution,
adopted several years ago in large fields, increases the risk of
resistance phenomena but does allow a substantial reduction in chemical
treatments, subject, however, to control of the proliferation of pests
until then of secondary importance. Attention today focuses on the
spatio-temporal aspects of the management of both harmful and useful
populations, in particular by devoloping and managing their vegetation
habitats. Accordingly, the consistency of agroecological actions by
agronomists and crop protection specialists is decisive: preventive
solutions are essential, at the scale of entire fields and at the scale
of larger land units, including uncultivated areas. Priority must be
accorded to balanced and sustainable functioning of agroecosystems. The
adoption of this new strategy by practitioners involves an important
break with traditional techniques and requires a marked change in
mentalities.

Document Number: 312

Biotechnology-based tools are now widely used to enhance and expand the
traditional remit of potato in food production. By modifying its
functionality, the capacity of the potato to produce, for example,
therapeutic or industrial compounds is now a reality, and its ability to
resist disease can also be radically improved. Two developments have
been crucial to expanding the role of potato: the recent advances in the
fields of structural and functional potato genomics and the ability to
integrate genes of interest into the potato genome. In this review we
discuss how both developments have diversified the remit of this crop.

Document Number: 313

In contrast to the dominating unsaturated C-18 fatty acid, medium-chain
fatty acids (MCFA) are nearly absent in the oil of common canola.
Modification of canola oil towards higher contents of C-8 to C-14 fatty
acids would create new possibilities for oleochemical usages, in both
the nutritional and the non-food sectors. For this purpose, spring
oilseed rape (cv. 'Drakkar') was genetically modified by introduction of
MCFA-encoding genes from Cuphea species containing approximately 90%
MCFA in their Two different single constructs involving the
3-ketoacyl-acyl carrier protein seed oil. synthase (KAS)III from C.
lanceolata were used, one harbouring the wild-type gene, C/KASIIIbwt,
and the other containing the point-mutated gene, C/KASIIIbmut, along
with two double constructs containing C/KASIIIbmut in combination with a
medium-chain thioesterase gene from C. lanceolata (C/FatB3) or C.
hookeriana (ChFatB2). For both single-gene constructs, a phenotype with
an increased content of MCFA was not detected; however, the C/KASIIIbwt
transformants produced up to 6.7% palmitic acid (C-16). In T2 seeds
bearing the C/KASIIIbmut/C/FatB3 double-gene construct, contents of up
to 2.9% capric (C-10) and 11.4% palmitic acid were achieved. The best
transformant with the gene construct C1KASIIIbmut/ChFatB2 contained 1.4%
caprylic acid (C-8) and 7.9% C-10, and these results were confirmed in
T3 seeds.

Document Number: 314

The current procedures to assess the safety of food and feed derived
from modern biotechnology include the investigation of possible
unintended effects. To improve the probability of detecting unintended
effects, profiling techniques such as proteomics are currently tested as
complementary analytical tools to the existing safety assessment. An
optimized two-dimensional gel electrophoresis (2DE) method was used as a
proteomics approach to investigate insertional and pleiotropic effects
on the proteome due to genetic engineering. Twelve transgenic
Arabidopsis thaliana lines were analyzed by 2DE, and their seed
proteomes were compared to that of their parental line as well as to 12
Arabidopsis ecotype lines. The genetic modification of the Arabidopsis
lines, using three different genes and three different promoters, did
not cause unintended changes to the analyzed seed proteome. Differences
in spot quantity between transgenic and nontransgenic lines fell in the
range of values found in the 12 Arabidopsis ecotype lines or were
related to the introduced gene.

Document Number: 315

Plant genetic engineering has the potential to introduce new allergenic
proteins into foods but, at the same time, it can be used to remove
established allergens. Here, we report the molecular characterization of
Lyc e 3, a new tomato (Lycopersicon esculentum) allergen, and the
efficient down-regulation of its expression in transgenic tomato plants.
Following the identification of an immunoglobulin E (IgE)-binding 9-kDa
polypeptide in tomato peel, designated Lyc e 3, its partial amino acid
sequence was determined by N-terminal protein sequencing. Sequence
comparison revealed that Lyc e 3 encodes a nonspecific lipid transfer
protein (ns-LTP). In plants, ns-LTPs are encoded by large gene families
which differ in primary amino acid sequence, expression and proposed
cellular function. To identify Lyc e 3 encoding complementary DNAs
(cDNAs), public tomato expressed sequence tag (EST) databases were
screened for ns-LTP sequences. Following this strategy, two cDNAs, LTPG1
and LTPG2, with high homology to the N-terminal sequence of Lyc e 3,
were identified. Ectopic expression of LTPG1 and LTPG2 in Escherichia
coli, followed by immunoblotting, verified their IgE reactivity.
Subsequently, transgenic tomato plants constitutively expressing LTPG1-
or LTPG2-specific double-stranded RNA interference (dsRNAi) constructs
were created and tested for the suppression of Lyc e 3 accumulation.
Efficient silencing of Lyc e 3 was documented by Northern and Western
blotting. In both cases, Lyc e 3 accumulation was decreased to levels
below the detection limit (less than 0.5% of the wild-type protein). The
allergenic potential of Lyc e 3-deficient tomato fruits was tested by
measuring histamine release from sensitized human basophils stimulated
with transgenic and parental lines. These assays revealed a strong (10-
to 100-fold) decrease in histamine release of human basophils challenged
with transgenic fruit extracts when compared with control extracts.
These results demonstrate the feasibility of creating low allergenic
tomato fruits by means of dsRNAi inhibition.

Document Number: 316

Three natural somatic mutations at codon 304 of the phytoene desaturase
gene (pds) of Hydrilla verticillata (L. f. Royle) have been reported to
provide resistance to the herbicide fluridone. We substituted the
arginine 304 present in the wild-type H. verticillata phytoene
desaturase (PDS) with all 19 other natural amino acids and tested PDS
against fluridone. In in vitro assays, the threonine (Thr), cysteine
(Cys), alanine (Ala) and glutamine (Gln) mutations imparted the highest
resistance to fluridone. Thr, the three natural mutations [Cys, serine
(Ser), histidine (His)] and the wild-type PDS protein were tested in
vitro against seven inhibitors of PDS representing several classes of
herbicides. These mutations conferred cross-resistance to norflurazon
and overall negative cross-resistance to beflubutamid, picolinafen and
diflufenican. The T3 generation of transgenic Arabidopsis thaliana
plants harbouring the four selected mutations and wild-type pds had
similar patterns of cross-resistance to the herbicides as observed in
the in vitro assays. The Thr304 Hydrilla pds mutant proved to be an
excellent marker for the selection of transgenic plants. Seedlings
harbouring Thr304 pds had a maximum resistance to sensitivity (R/S)
ratio of 57 and 14 times higher than that of the wild-type for
treatments with norflurazon and fluridone, respectively. These plants
exhibited normal growth and development, even after long-term exposure
to herbicide. As Thr304 pds is of plant origin, it could become more
acceptable than other selectable markers for use in genetically modified
food.

Document Number: 317

The yield potential (Y(p)) of a grain crop is the seed mass per unit
ground area obtained under optimum growing conditions without weeds,
pests and diseases. It is determined by the product of the available
light energy and by the genetically determined properties: efficiency of
light capture (epsilon(i)), the efficiency of conversion of the
intercepted light into biomass (epsilon(c)) and the proportion of
biomass partitioned into grain (eta). Plant breeding brings eta and
epsilon(i) close to their theoretical maxima, leaving epsilon(c),
primarily determined by photosynthesis, as the only remaining major
prospect for improving Y(p). Leaf photosynthetic rate, however, is
poorly correlated with yield when different genotypes of a crop species
are compared. This led to the viewpoint that improvement of leaf
photosynthesis has little value for improving Y(p). By contrast, the
many recent experiments that compare the growth of a genotype in current
and future projected elevated [CO(2)] environments show that increase in
leaf photosynthesis is closely associated with similar increases in
yield. Are there opportunities to achieve similar increases by genetic
manipulation? Six potential routes of increasing epsilon(c) by improving
photosynthetic efficiency were explored, ranging from altered canopy
architecture to improved regeneration of the acceptor molecule for
CO(2). Collectively, these changes could improve epsilon(c) and,
therefore, Y(p) by c. 50%. Because some changes could be achieved by
transgenic technology, the time of the development of commercial
cultivars could be considerably less than by conventional breeding and
potentially, within 10-15 years.

Document Number: 318

The application of a ligation-dependent probe amplification (LPA)
technique to the simultaneous event-specific detection and relative
quantification of DNA from genetically modified organisms in foods is
described. The system is based on the ligation of synthetic bipartite
probes when hybridized to the corresponding target DNA sequence. The
ligation products possess lengths characteristic for each target
sequence. Universal primer binding sites (PBS) at the 5' and 3'-ends
enable their subsequent competitive amplification using one common pair
of primers. The use of one fluorescein (FAM) labeled primer permits
amplification products to be separated and detected via capillary
electrophoresis. Respective probes were designed to allow the detection
of reference genes in the genomes from maize and soya, as well as of
event-specific junction regions in the transgenic maize line MON810 and
in Roundup Ready soya. Specificity, sensitivity, and the potential of
the technique for the relative quantification of recombinant DNA were
assayed using mixtures of DNA extracted from certified reference maize
and soybean flours. The novel strategy results in a modular system which
can be complemented by further probes to broaden the range of target
sequences.

Document Number: 319

With the rapid development of the transgenic technique, more and more
transgenic food, especially involved in soybean oil, has been found in
the market, it has attracted much attention in terms of transgenic
food's edible safety. The content of 9 kinds of heavy metals was
analyzed in transgenic soybean oils. The average recoveries for T, Cr,
Mn, Cu, Zn, As, Cd, Pb, Hg was achieved, which ranged from 96 percent to
105.78 percentt, relative standard derivation (RSD) 0.98 percent similar
to 4.08 percent (n=5). The results showed that contents of 9 kinds of
heavy metals ranged from 0.00 ng/g to 930.00ng/g. The first three heavy
metals was ranked by Zn > Cr > T, particularly the content of Zn was
over 700ng/g, even more than 930ng/g. The content of Pb is less than
100ng/g, and Cd near to zero. The data analysis showed that transgenic
soybean oil in Beijing was presumably polluted with Zn, Cr and Pb,
further investigation over a range of sites in Beijing will be conducted
in future, to make more clear conclusions.

Document Number: 320

Major staple crops are often deficient in some of the nutrients required
in human diet. Thus, malnutrition is a major problem, especially in
developing countries, where a diversified diet is not affordable for the
majority. Several strategies have been adopted to improve nutrition.
However, micronutrient deficiency is still widely spread. Rice is one of
the most important staple foods for a large part of the world's
population. Therefore, even a small improvement in nutritional content
of rice seeds can have a dramatic impact on human health. Different
approaches are being exploited to produce rice enhanced in nutrients
with iron and provitamin A.

Document Number: 321

Carotenoids participate in light harvesting and are essential for
photoprotection in photosynthetic plant tissues. They also furnish
non-photosynthetic flowers and fruits with yellow to red colors to
attract animals for pollination and dispersal of seeds. Although animals
can not synthesize carotenoids de novo, carotenoid-derived products such
as retinoids (including vitamin A) are required as visual pigments and
signaling molecules. Dietary carotenoids also provide health benefits
based on their antioxidant properties. The main pathway for carotenoid
biosynthesis in plants and microorganisms has been virtually elucidated
in recent years, and some of the identified biosynthetic genes have been
successfully used in metabolic engineering approaches to overproduce
carotenoids of interest in plants. Alternative approaches that enhance
the metabolic flux to carotenoids by upregulating the production of
their isoprenoid precursors or interfere with light-mediated regulation
of carotenogenesis have been recently shown to result in increased
carotenoid levels. Despite spectacular achievements in the metabolic
engineering of plant carotenogenesis, much work is still ahead to better
understand the regulation of carotenoid biosynthesis and accumulation in
plant cells. New genetic and genomic approaches are now in progress to
identify regulatory factors that might significantly contribute to
improve the nutritional value of plant-derived foods by increasing their
carotenoid levels.

Document Number: 322

In this review, we describe the need and progress to improve the iron
and zinc contents in crop plants by genetic means. To achieve this goal
either by transgenic approaches or classical breeding, knowledge about
the physiological and molecular mechanisms of mineral uptake and mineral
homeostasis will be very helpful. The progress in our understanding of
the molecular processes and genes is described, and the use of the
identified genes by transgenic approaches is illustrated. Genetic
mapping of the existing variation will allow marker-assisted breeding to
exploit the available natural variation in crop plants. For this
application, ultimately the knowledge of the genes underlying this
quantitative variation, called quantitative trait loci (QTL), will be
required. It is expected that research in this field in the model
species Arabidopsis thaliana, where the molecular tools are available,
might help in the identification of the allelic variation at QTL.

Document Number: 323

Several countries have introduced mandatory labeling requirements on
foods derived from genetically modified organisms. Real-time
quantitative Polymerase Chain Reaction (PCR) has quickly become the
method of choice in support of these regulations and requires the
development of separate PCR assays targeting the transgenic sequence as
well as a specific endogenous gene sequence. To develop a Brassica
napus-specific PCR assay, partial sequences of the acetyl-CoA
carboxylase BnACCg8 gene from B. napus and the closely related Brassica
rapa were determined and compared, and a region of unique nucleoticle
sequence was identified. Universal amplification primers were designed
to either side of this region, and a locked nucleic acid TaqMan probe
was designed to the B. napus-specific sequence. Evaluation of this
primer/probe combination indicated a high level of specificity to B.
napus: no amplification signal was observed with any other species
tested, including five closely related Brassica species. The method was
assayed with 14 different B. napus cultivars, and comparable
amplification curves were consistently obtained for all. The assay was
highly sensitive, with a limit of detection between 1 and 10 haploid
copies. Practically, the method was demonstrated to be effective for the
detection of processed food samples and for the quantification of
Roundup Ready canola content in mixed samples.

Document Number: 324

The potential for applying biotechnology to benefit animal agriculture
and food production has long been speculated. The addition of human milk
components with intrinsic antimicrobial activity and positive charge to
livestock milk by genetic engineering has the potential to benefit
animal health, as well as food safety and production. We generated one
line of transgenic goats as a model for the dairy cow designed to
express human lysozyme in the mammary gland. Here we report the
characterization of the milk from 5 transgenic females of this line
expressing human lysozyme in their milk at 270 mu g/mL or 68% of the
level found in human milk. Milk from transgenic animals had a lower
somatic cell count, but the overall component composition of the milk
and milk production were not different from controls. Milk from
transgenic animals had a shorter rennet clotting time and increased curd
strength. Milk of such nature may be of benefit to the producer by
influencing udder health and milk processing.

Document Number: 325

The paper examines the reasons for and consequences of lost domestic
animal biodiversity. The origin of domestic poultry and livestock
diversity is reviewed from the first center of domestication in the
Middle East during the Neolithic Revolution. Accompanied by domestic
animals and birds, mankind spread worldwide over the last 12,000 yr,
thereby increasing domestic animal biodiversity via adaptation to many
environmental challenges, resulting in about 6,000 breeds within only a
small number of species used for food. During the last 50 yr of the 20th
century, about 20% of these livestock and poultry breeds have become
extinct, and the remainder is at risk. This erosion of unique
biodiversity is due to changes in farm practices developed in the West
that involve mono-breed, intensive farming systems that are
unsustainable. The close symbiotic relationship of Homo sapiens and
domestic animals and birds over millennia is changing, resulting in a
lost understanding of sustainability among urban communities. The
single-minded focus on profit is resulting in the loss of the historic
European and Western culture based on Judeo-Christian values. Respect
for biological boundaries, community, and quality of life are
disappearing in Western society. Concurrently, farming is now only a
business. The principal decision makers are no longer farmers but
business executives, who are remote from the farm. The emphasis on cheap
food is the principal driver that leads to increased competition and
unsustainable practices. Farmers as well as their breeds are
disappearing. The advent of gene technology and transgenic livestock is
reviewed with the prospect of extensive manipulation of animal form and
function and abuse of genotypes as animals are redesigned, suffer, and
lose all dignity. By handling its animals in this manner, high Western
civilization is losing its culture and values and becoming simply the
top animal species by using its power selfishly. The case is presented
that the extinction of animals indicates an urgent need to conserve
historic Western culture before it, too, is lost.

Document Number: 326

Evidence continues to mount that dietary components are important
determinants of cancer risk and tumor behavior Although these linkages
are fascinating, numerous inconsistencies are also evident in the
literature. Although multifactorial, these discrepancies likely reflect
variation in the ability of food constituents to reach and/or modify
critical molecular targets. Genetic polymorphisms can alter the response
to dietary components (nutrigenetic effect) by influencing the
absorption, metabolism, or site of action. Likewise, variation in DNA
methylation patterns and other epigenomic events that influence overall
gene expression can influence the biological response to food components
and vice versa. Fluctuations in the ability of food components to
increase or depress gene expression (nutritional transcriptomic effect)
may also account for some of the inconsistencies in the response to
foods. Functional proteomic studies that capture all of the proteins
produced by a species and link them to physiological significance within
the cell will be fundamental to understanding the relationship between
dietary interventions, proteome changes, and cancer Although a bioactive
food component may influence a number of key molecular events that are
involved with cancer prevention, to do so it must achieve an effective
concentration within the target site, be in the correct metabolic form,
and bring about a change in one or more small molecular weight signals
in the cellular milleau (metabolomic effects). Fundamental to assessing
and evaluating the significance of the interrelationships among
bioactive food components with nutrigenetics, nutritional epigenomics,
nutritional transcriptomics, proteomics, and metabolomics is knowledge
about the appropriate tissue/cell or surrogate to evaluate and validated
biomarkers that reflect changes in each. As the era of molecular
nutrition grows, a greater understanding about the role of foods and
their components on cancer risk and tumor behavior will surely unfold.
Such information will be critical in the development of effective
preemptive approaches to reduce the cancer burden.

Document Number: 327

As a step towards developing an efficient genetically modified organism
(GMO) detection system, the present investigation proposes microarray as
a prototype for high-throughput detection of pure GMO samples. Common
T-DNA regions of the expression cassette such as 35S promoter, 35S
terminator, nopaline synthase terminator (NOSt), hygromycin, an
kanamycin selection marker genes were detected in the transgenic rice,
tomato, and potato developed in our laboratory. For proof-of-concept
purposes, market-introduced GM potato, GM maize, and cornflakes were
screened. The non-GM potato and maize control detected only the presence
of endogenous genes while the GM targets detected the presence of
transgenic genes such as CaMV 35S promoter, 35S terminator, NOSt and (n)
over bar ptII, pat, cp4 epsps, cry1 Ab genes on chips. Moreover, it was
observed that the sensitivity of this system for serially diluted GM
potato tubers was up to 0.01-0.002% of the mass fraction. Due to its
high accuracy and speed, it is believed that the microarray detection
system will play an important role in routine, high-throughput detection
of pure GMO samples in the future.

Document Number: 328

Different foodstuffs for humans and monogastric animals were analysed to
detect CryIA (b) gene and quantify the CryIA(b) protein present in the
transgenic maize used as an ingredient. Eight out of 32 foods obtained
from the market showed to have been elaborated with transgenic Bt maize.
Specific primers used to identify the transgenic event revealed that
Mon810 was predominantly present in the foodstuffs. A commercial ELISA
test allowed the quantification of the CryIA(b) protein in low processed
foods, and found that 0.1 ppm was the highest value per gram of food. A
Western blot carried out with immuno-purified polyclonal antibodies was
capable of detecting both the intact or degraded CryIA(b) protein
depending on the food assayed. (c) 2005 Elsevier Ltd. All rights
reserved.

Document Number: 329

Real Time PCR is a technique widely used for GMO quantification. The
maize seed contains diploid embryo and triploid endosperm. This implies
that the ratio transgene/endogene determined by Real Time PCR will vary
due to the presence of different genome copies in the seed compartments.
The transgene/endogene ratio is determined for single seeds from
commercial transgenic line T25. This ratio is compared between the seeds
that originate from reciprocal crosses between T25 and the conventional
variety B73. The variation in the GMO content due to the presence of
seeds with different origin in the seed samples of 3000 seeds is
evaluated.

Document Number: 330

Hepatitis B virus is responsible for significant morbidity and mortality
despite the availability of safe and effective injectable vaccines.
Transgenic plants are a novel system for expression and oral delivery of
subunit vaccine antigens. In this paper, plant expression vector pBIS2S,
which contains the HBsAg gene with precursor sequence preS2, was
reconstructed based on pBI121 and was introduced into tobacco by the
co-cultivating leaf-section method via Agrobacterium tumefaciens
LBA4404. After selection by kanamycin and confirmation by PCR and
Southern blot, it was found that the objective fragment had integrated
into the tobacco genome. ELISA analysis showed that the hepatitis B
surface antigen (HbsAg) was expressed in transgenic tobacco plants.
These results indicate that the production of HbsAg with PreS2 extended
to food plants as a source of edible vaccines is possible.

Document Number: 331

Insect pests are major problems for all crops, worldwide. In this review
we will focus on legumes, which are attacked by a range of insect pests
including pod/seed feeders, defoliators and sap feeders. We review the
history of breeding for resistance to insect pests in legumes, which has
had mixed success, and discuss further opportunities in this area. We
also review the extraordinary array of direct and indirect mechanisms
contributing to insect defence in legumes, the understanding and
exploitation of which offer opportunities for both legume and non-legume
crops. There is also good potential to improve insect resistance in
legume crops through a detailed understanding of the signaling pathways
that regulate induced responses to insect feeding, and recent progress
in this area, primarily obtained from non-legume systems, is reviewed.
The importance legumes play in farming systems, their wide range of
novel chemistry and the emergence of model systems suitable for genomic
approaches present opportunities for research in this area strongly
linked to breeding programs to help develop legume crops with enhanced
insect resistance.

Document Number: 332

Fruit ripening is a unique plant developmental process with direct
implications for our food supply, nutrition, and health. In contrast to
climacteric fruit, where ethylene is pivotal, the hormonal control of
ripening in nonclimacteric fruit, such as grape (Vitis vinifera), is
poorly understood. Brassinosteroids (BRs) are steroidal hormones,
essential for normal plant growth and development but not previously
implicated in the ripening of nonclimacteric fruit. Here we show that
increases in endogenous BR levels, but not indole-3-acetic acid (IAA) or
GA levels, are associated with ripening in grapes. Putative grape
homologs of genes encoding BR biosynthesis enzymes
(BRASSINOSTEROID-6-OXIDASE and DWARF1) and the BR receptor
(BRASSINOSTEROID INSENSITIVE 1) were isolated, and the function of the
grape BRASSINOSTEROID-6-OXIDASE gene was confirmed by transgenic
complementation of the tomato (Lycopersicon esculentum) extreme dwarf
(d(x)/d(x)) mutant. Expression analysis of these genes during berry
development revealed transcript accumulation patterns that were
consistent with a dramatic increase in endogenous BR levels observed at
the onset of fruit ripening. Furthermore, we show that application of
BRs to grape berries significantly promoted ripening, while
brassinazole, an inhibitor of BR biosynthesis, significantly delayed
fruit ripening. These results provide evidence that changes in
endogenous BR levels influence this key developmental process. This may
provide a significant insight into the mechanism controlling ripening in
grapes, which has direct implications for the logistics of grape
production and down-stream processing.

Document Number: 333

Starch is a readily renewable resource that is very widely used for food
and industrial purposes; however, greater variation in the functional
properties of starch would further extend the use of this biodegradable
polymer. Genetic engineering may provide a way to produce designer
starches that have the desired properties. Starch-binding domains (SBD)
from bacterial enzymes that catabolise starches have the ability to bind
two helices of starch and thus have the potential to crosslink starch
and/or to be used as anchors for other enzymes that can modify starch
properties. In a first step towards novel modi. cation of starch we have
investigated the effect of expressing SBDs, singly and in tandem, in
planta, and targeting them to the chloroplast in the model plant
Arabidopsis thaliana ( L.) Heynh. Transgenic plants that contained the
SBD from the cyclomaltodextrin glucanotransferase (CGTase) of
Thermoanaerobacterium thermosulfurigenes in the chloroplast were
produced in both the wild type and the starch excess mutant (sex 1-1)
backgrounds. Analysis of starch isolated from the chloroplasts of these
lines revealed no significant changes in the amylose : amylopectin
ratio, the chain-length distribution of debranched amylopectin or the
gelatinisation temperature when compared to the parental line. However,
significant changes were observed in the starch granule size with the
plants expressing the construct having larger granules. The effect was
more pronounced in the sex 1-1 background, and expression of two
starch-binding domains linked in tandem had an even greater effect.
Despite the starch granules being larger in lines expressing the
starch-binding domain, no difference was seen in the starch content of
the leaves when compared to parental lines. As the presence of the SBDs
in the starch granule only altered granule size, and not other granule
properties, they may provide an ideal anchor for targeting
starch-modifying enzymes to the site of starch synthesis. This will
allow the development of novel modi. cations of starch during synthesis.

Document Number: 334

Roundup-Ready soybeans have been genetically modified to resist the
effects of the herbicidal glyphosate and have become the most prevalent
transgenic crop in the world. In this work, Roundup-Ready soybeans were
used as raw material to study the effects of critical processing
procedures such as grinding, cooking, blending, homogenization,
sterilization, and spray-drying on the length of DNA fragments of an
endogenous gene (lectin) and an exogenous gene (epsps) examined in
material from three soybean foods of bean curd, soy milk, and soy powder
and from samples taken during their processing. The results showed that
various processing procedures caused degradations of both the endogenous
and exogenous genes to different degrees. In the grinding procedure,
endogenous gene DNA was degraded from 1883 to similar to 836 bp, and
exogenous gene DNA was degraded from 1512 to similar to 408 bp. In the
blending and squeeze-molding procedures, exogenous gene DNA was also
degraded from about 408 to 190 bp, but there was no obvious action on
the endogenous gene. After the endogenous and exogenous genes had been
degraded to some degree, such as 836 and 408 bp, respectively, they were
not evidently affected by cooking procedure at 100 degrees C for 15 min.
However, the endogenous gene was further considerably degraded from
around 836 to 162 bp in the sterilization procedure at 121 degrees C for
30 s. The effect of the homogenization step on endogenous and exogenous
genes was similar to that of the cooking procedure. The coagulation
procedure, principally a biochemical reaction, did not greatly affect
the exogenous gene but did affect endogenous gene, reducing DNA size
from about 836 to 407 bp. Furthermore, the spray-drying procedure, a
process of physical shearing, high temperature, and sudden high
pressure, distinctly caused degradation of both the lectin and epsps
genes, rapidly decreasing the sizes from about 836 to 162 bp for the
endogenous gene and from about 408 to 190 bp for the exogenous gene.

Document Number: 335

Milling fractions from conventional and transgenic corn were prepared at
laboratory scale and used to study the influence of sample composition
and heat-induced DNA degradation on the relative quantification of
genetically modified organisms (GMO) in food products. Particle size
distributions of the obtained fractions (coarse grits, regular grits,
meal, and flour) were characterized using a laser diffraction system.
The application of two DNA isolation protocols revealed a strong
correlation between the degree of comminution of the milling fractions
and the DNA yield in the extracts. Mixtures of milling fractions from
conventional and transgenic material (1%) were prepared and analyzed via
real-time polymerase chain reaction. Accurate quantification of the
adjusted GMO content was only possible in mixtures containing
conventional and transgenic material in the form of analogous milling
fractions, whereas mixtures of fractions exhibiting different particle
size distributions delivered significantly over- and underestimated GMO
contents depending on their compositions. The process of heat-induced
nucleic acid degradation was followed by applying two established
quantitative assays showing differences between the lengths of the
recombinant and reference target sequences (A, Delta/(A) = -25 bp; B,
Delta/(B) = +16 bp; values related to the amplicon length of the
reference gene). Data obtained by the application of method A resulted
in underestimated recoveries of GMO contents in the samples of
heat-treated products, reflecting the favored degradation of the longer
target sequence used for the detection of the transgene. In contrast,
data yielded by the application of method B resulted in increasingly
overestimated recoveries of GMO contents. The results show how commonly
used food technological processes may lead to distortions in the results
of quantitative GMO analyses.

Document Number: 336

Inulin- type fructans are stored in the tuberous roots of the Brazilian
cerrado plant Viguiera discolor Baker ( Asteraceae). In Cynara scolymus
( artichoke) and Echinops ritro ( globe thistle), the fructans have a
considerably higher degree of polymerization ( DP) than in Cichorium
intybus ( chicory) and Helianthus tuberosus ( Jerusalem artichoke). It
was shown before that the higher DP in some species can be attributed to
the properties of their fructan: fructan 1- fructosyl transferases ( 1-
FFTs; EC 2.4.1.100), enzymes responsible for chain elongation. Here, we
describe the cloning of a high DP ( hDP) 1- FFT cDNA from V. discolor
and its heterologous expression in Pichia pastoris. Starting from 1-
kestose and Neosugar P ( a mixture of oligo- inulins from microbial
origin) as substrates, the recombinant enzyme produces a typical hDP
inulin profile in vitro, closely resembling the one observed in vivo.
The enzyme shows no invertase activity and sucrose: sucrose 1- fructosyl
transferase ( 1- SST; EC 2.4.1.99) activity in vitro. Pattern evolution
during incubation suggests that inulins with DP >= 6 are much better
substrates than sucrose or lower DP oligo- fructans. Because hDP inulin-
type fructans show superior properties for specific food and non- food
applications, the hDP 1- FFT gene from V. discolor has potential for the
production of hDP inulin in vitro or in transgenic crops.

Document Number: 337

Possibilities and limits of the detection of genetically modified (gm)
rape in rape honey by Real-time PCR are described. Based on a published
method for DNA extraction from honey the sensitivity of PCR methods for
the detection of gm rape is demonstrated. Honeys with high numbers of
rape pollen resulted in high yields of amplifiable rape DNA. Low
sensitivity was observed in honeys with rape pollen below 50% of the
total pollen fraction. The amount of amplifiable transgenic DNA can
indicate the proportion of gm rape pollen to total pollen number in rape
honeys. In the future, sensitive methods for the analysis of transgenic
rape in honey will not only be significant for food analysis. They can
also be used as a tool for monitoring programs of field releases or
coexistence studies of transgenic and conventional plants.

Document Number: 338

Humans require more than 22 mineral elements, which can all be supplied
by an appropriate diet. However, the diets of populations subsisting on
cereals, or inhabiting regions where soil mineral imbalances occur,
often lack Fe, Zn, Ca, Mg, Cu, I or Se. Traditional strategies to
deliver these minerals to susceptible populations have relied on
supplementation or food fortification programs. Unfortunately, these
interventions have not always been successful. An alternative solution
is to increase mineral concentrations in edible crops. This is termed
'biofortification'. It can be achieved by mineral fertilization or plant
breeding. There is considerable genetic variation in crop species that
can be harnessed for sustainable biofortification strategies. Varieties
with increased mineral concentrations in their edible portions are
already available, and new genotypes with higher mineral densities are
being developed.

Document Number: 339

Serious attempts to estimate the impact of allergic reactions to foods
on public health did not begin until the 1980s. Until about 15 years ago
food allergy was considered a minor aspect of food safety. Two
developments probably prompted a radical re-appraisal of that situation.
The first was the apparently inexorable rise in the prevalence of atopic
diseases, of which food allergy forms a part, with its possible
consequences highlighted by some well-publicised severe reactions. The
second was the growth of genetic modification technology, manifested by
the commercialisation of transgenic crops. Each of these developments
impacted on the food industry in distinct ways. On the one hand,
consumers with food allergies had to be enabled to avoid specific
allergens in products formulated with existing ingredients. Food
manufacturers therefore had to identify those specific allergens down to
trace amounts in all the ingredients forming the product and label or
remove them. On the other hand, the introduction of products using
ingredients from novel sources required an assessment of the
allergenicity of these ingredients as an integral part of safety
assurance. The approaches used by the food industry to protect existing
consumers who have food allergies and those at potential risk of
sensitisation from novel proteins will be illustrated, emphasising how
they need to be built into every stage of the life cycle of a product.

Document Number: 340

Since genetically modified ( GM) crops and foods began to appear in
market, the detection of these GM foods has become an important issue.
Efficient and reliable methods are urgently needed to analyze the GM
content of a large quantity of foods. However, most foods are processed
through heating or cooking, and their proteins are denatured. Therefore,
DNA rather than protein is the major target for detecting GM foods. In
this report, polymerase chain reaction (PCR) was performed with dual
sets of DNA primers, which co-amplified the soybean-specific lectin gene
and the transgenic petunia transit peptide sequence (CTP) in the 5' end
of the 5-enol-pyruvyl-shikimate-3-phosphate synthase (EPSPS) gene within
the same reaction. PCR with these two sets of primers reacted
specifically with lectin gene and CTP, and can detect soybean with GM
content less than 1%. PCR detecting both lectin gene and CTP was also
applied to a soybean-derived product tou-kan, a traditional oriental
food. Results indicated that although DNA was partially degraded in
tou-kan, this PCR method was still able to detect the presence of EPSPS
gene. However, when the DNA within tou-kan was destroyed badly, neither
lectin nor EPSPS genes were detected by the PCR suggested here. Finally,
an examination procedure for the Roundup Ready soybean was suggested
according to the results of PCR with dual pairs of DNA primers. The
proposed PCR method has proved to be a reliable and efficient method for
detecting the GM content of the processed foods from Roundup Ready
soybean.

Document Number: 341

There is growing scientific evidence that consumption of n-3 very
long-chain polyunsaturated fatty acids (n-3 VLC-PUFA) helps in brain and
eye development, and protects against a range of common degenerative
diseases. This has provided the impetus to the scientists to develop new
and renewable sources for these important fatty acids so that the food
industry is able to produce and market products fortified with n-3
VLC-PUFA. The bioactive efficacy and stability of food products
containing n-3 VLC-PUFA may be determined not only by the amount of n-3
VLC-PUFA present but also by the positional distribution of these acids
within the triacylglycerol (TAG) molecules (regiopurity). Studies of the
effects of positional distribution on the functionality of n-3 VLC-PUFA
containing oils have been hampered by a general lack of pure TAG
regioisomers for experimentation. This paper reviews methods that have
been used for the synthesis of TAG regioisomers containing n-3 VLC-PUFA,
with special reference to those in which one n-3 VLC-PUFA occurs in
combination with two long-chain saturated acids.

Document Number: 342

Genetically modified (GM) tomatoes have been approved for
commercialization in many countries since the first GM tomato FLAVR SAVR
was permitted for planting in 1994. In China, GM tomato Huafan No I with
a character of long shelf-life was the first GM plant which was approved
for commercialization in 1996. To meet the requirement of the GM
tomatoes labeling policy that has been actualized in China since 2001,
screening and construct-specific PCR detection methods for detecting the
universal elements transformed into tomato, such as cauliflower mosaic
virus 35S (CaMV35s) promoter and the nopaline synthase (NOS) terminator
of Agrobacterium tumefaciens, and the specifically inserted heterologous
DNA sequence between CaMV35s promoter and anti-sense ethylene-forming
enzyme (EFE) gene were set up. To make the detection methods normative,
a novel single copy tomato gene LAT52 was also used as an endogenous
reference gene in the PCR detection systems. The limit of detection of
screening and construct specific detection methods for Huafan No 1 was
68 haploid genome copies in conventional PCR detection, and three copies
in TaqMan real-time PCR detection. The limit of quantitation of
screening quantitative PCR assays for Huafan No 1 was three copies and
was 25 copies for construct-specific quantitative PCR. Two samples with
known Huafan No I tomato content were detected using the established
conventional and real-time PCR systems, and these results also indicated
that the established Huafan No I screening and construct-specific PCR
detection systems were reliable, sensitive and accurate. (c) 2005
Society of Chemical Industry.

Document Number: 343

The objectives of this study were to use transgenic sows that
overexpress IGF-I in milk to investigate the effect of a short-term fast
on piglet intestinal morphology and disaccharidase activity and to
determine how milk-borne IGF-I influences the response to fasting. After
farrowing, litters were normalized to 10 piglets. On d 6, piglets (n =
30) suckling IGF-I transgenic (TG) sows and piglets (n = 30) suckling
non-transgenic sows (control) were assigned randomly to three
treatments: fed piglets (0 h), which remained with the sow until
euthanized on d 7, or fasted piglets, which were removed from the sow at
either 6 or 12 h before euthanasia on d 7. Serum IGF-I and IGFBP,
intestinal weight and length, jejunal protein and DNA content,
disaccharidase activity, and villus morphology were measured. Fasting
for 12 h resulted in a negative weight change between d 6 and 7
(quadratic response to fasting; P < 0.001). Piglets suckling TG sows
tended to have greater intestinal length (P = 0.068), but no effect of
IGF-I overexpression was noted for intestinal weight. Fasting, however,
resulted in linear (P < 0.001) and quadratic (P = 0.002) decreases in
intestinal weight. Serum IGF-I did not differ between control and TG
sows, but decreased linearly (P = 0.003) with fasting. Serum IGFBP-4
decreased (linear and quadratic; P:5 0.02) with fasting, whereas IGFBP-1
increased quadratically (P < 0.001) with fasting. Jejunal villus height,
width, and crypt depth were all increased with fasting (linear and
quadratic; P < 0.04). Disaccharidase activity was not affected by fed
state; however, piglets suckling TG sows had greater jejunal
lactase-phlorhizin hydrolase (P < 0.01) and sucrase-isomaltase (P =
0.02) activities than control piglets. In summary, intestinal weight,
villus morphology, serum IGF-1, serum IGFBP1 and -4, and piglet BW
change were altered (P < 0.02) in response to fasting. Thus, the
duration of food deprivation before euthanization should be considered
when designing experiments to assess intestinal development or the IGF
axis, as the magnitude of differences between the fed and fasted state
may exceed those expected as a result of experimental treatment.

Document Number: 344

Molecular studies have shown that several genes with various functions
are induced by environmental stresses such as drought, high-salinity and
low temperature in plants. Most of the dehydration responsive genes are
induced by the plant hormone abscisic acid (ABA), but others are not.
Expression analyses of dehydration-responsive genes have provided at
least four independent regulatory systems (regulons) for gene expression
in a model plant Arabidopsis thaliana. The cis-acting elements in the
promoters of some genes that have a typical stress-inducible expression
profile and the transcription factors that affect the expression of
these genes have been analyzed. Transcription factors that bind to a
DRE/CRT (dehydration-responsive element/C-repeat) cis-acting element
were isolated and termed DREB1/CBF (DRE-binding protein I/C-repeat
binding factor) and DREB2 (DRE-inding protein 2). Overexpression of
DREB1/CBF in transgenic Arabidopsis plants increased tolerance to
freezing, drought and high salt concentrations. The DREB1/CBF genes have
been successfully used to improve abiotic stress tolerance in a number
of different crop plants. Studies on the other transcription factors
associated with stress response are in progress. We collaborate with
many research groups to improve stress tolerant crop plants utilizing
regulon biotechnology. We hope the results of these collaborative
studies will contribute to the sustainable food production in developing
countries and help to prevent the global-scale environmental damage.

Document Number: 345

When herbivores feed on insect-resistant transgenic plants, food
consumption and utilization are probably affected. Quantifying these
parameters is of importance, because they may be associated with the
performance of parasitoids, which are important antagonists of
herbivorous pests. In this study, relative consumption rate (RCR),
relative growth rate (RGR), approximate digestibility (AD), efficiency
of conversion of digested food into body matter (ECD) and efficiency of
conversion of ingested food into body matter (ECI) of Chilo partellus
Swinhoe larvae (Lepidoptera: Crambidae) exposed to different
concentrations of transgenic insect-resistant Bacillus thuringiensis-com
(Bt-corn) tissue suspension, were investigated under laboratory
conditions. For the control groups, different concentrations of control
corn tissue suspension were used. In order to determine whether Bt
effects were influenced by larval age, 4- and 17-day-old larvae were
used. At all concentrations and for larvae of both ages, RGR, RCR, ECD
and ECI values in the control were significantly higher than in the Bt
group. Moreover, there was a gradual decrease in RCR, RGR and ECI values
when Bt concentrations were increased. However, AD values at the lowest
Bt concentration were significantly higher than in the control, whereas
no significant differences between both groups could be detected at
higher concentrations. In conclusion, the experiments showed that
nutritional indices of herbivores exposed to transgenic insect-resistant
plants could be quantified to determine the effects of these plants on
target or non-target organisms. This could be one component in future
evaluation of the effect of transgenic insect-resistant plants on the
second and third trophic level.

Document Number: 346

Qualitative and quantitative polymerase-chain-reaction-based methods to
detect genetically modified (gm) soy (Roundup-Ready (TM) soy) and maize
(Bt176 Maximizer maize; Bt11 maize, MON810 Yield Gard corn, T25
Liberty(R) Link maize) were applied to processed foods sold commercially
in Brazil. In total 100 foods containing maize and 100 foods containing
soy were analysed in 2000 and again in 2001. In 2000, 13% of the soy
containing products and 8% of those containing maize were shown to
consist of material derived from the corresponding genetically modified
organisms. In five samples of the products containing gm soy < 4% and in
eight samples > 4% of the soy derived material was identified as coming
from gm soybeans, whereas in the case of products containing gm maize
five samples of the maize derived material were found to contain < 4%
and three samples > 4% of gm maize. In 2001, gm soy was found in 21% of
the soy containing foods, and gm maize was found in 9% of the maize
containing products. Eight samples of the products containing gm soy
were shown to contain < 4% and 13 samples > 4% of gm soy, the
corresponding values for gm maize were five samples < 4% and four
samples > 4%. (c) 2004 Elsevier Ltd. 

Document Number: 347

Genetic engineering techniques have so far had limited impact in the
field of ornamental horticulture. As outlined in this review,
transformation systems and potential genes of interest are available. As
the development of new, novel varieties is an important driving force in
the industry, there are, therefore, good prospects for the development
of genetically modified ornamental varieties. The few products in the
market to date may simply be a reflection of the relatively small scale
of the industry compared to the major food crops, and the wide diversity
of species within it. Commercial issues attendant to the use of gene
technology in ornamental plants need careful consideration. These
include careful choice of crop and background variety, the international
regulatory process and freedom to operate.

Document Number: 348

Cassava is a poor man's crop which is mainly grown as a subsistence crop
in many developing countries. Its commercial use was first as animal
feed (also known as tapioca), but has shifted since the late sixties to
a source of native starch. The availability of native starches, which on
the one hand do not require substantial chemical derivatisation and on
the other hand have improved properties, would make cassava also for
small farmers a potentially attractive cash crop. Since breeding is
difficult in this polyploid, vegetatively propagated, crop a transgenic
approach would be ideal to improve certain characteristics. We have
created a cassava genotype producing amylose-free starch by genetic
modification. The absence of amylose increased the clarity and stability
of gels made with the transgenic starch, without requiring treatment
with environment-unfriendly chemicals such as epoxides (propylene oxide,
ethylene oxide) and acetic anhydride, which are normally used to improve
stability. The amylose-free starch showed no changes in particle size
distribution, chain length distribution or phosphorous content when
compared to amylose-containing starch, but the granule melting
temperature was increased by almost 2 degrees C. Furthermore, the
amylose-free cassava starch shows enhanced clarity and stability
properties. These improved functionalities are desired in technical
applications in paper and textile manufacturing, but also in the food
industry for the production of sauces, dairy products and noodles.

Document Number: 349

\During the last 20 years recombinant biotechnology has resulted in the
development of organisms with unique genetic compositions, some of which
are for intentional release to the environment. While concerns have been
raised that such organisms may be capable of inducing transient
unintended environmental effects, longer-term perturbations to soil
processes and non-target species effects have yet to be demonstrated. In
parallel with the growth of the commercial biotechnology industry has
come a significant growth in regulatory review processes intended to
evaluate the risks of these GMO products. Under the Toxic Substances
Control Act (TSCA), certain new microbial products that undergo
pre-manufacture review are examined for human and environmental risks
using data and other information received in accordance with the U.S.
Environmental Protection Agency's (EPA's) "Points to Consider" guidance
document. In the risk assessment process, carried out under the Federal
Insecticide, Fungicide, and Rodenticide Act (FIFRA) and the Federal
Food, Drug and Cosmetic Act (FFDCA) authorities, EPA evaluates both
microbial pesticide products and plants with pesticidal properties to
determine if Federal safety standards are met. For all pesticide
products, including genetically engineered pesticides, EPA receives
testing of product composition and chemical properties, human health
effects, environmental effects on non-target pests, and the fate of the
pesticide in the environment. The EPA's Office of Research and
Development supports risk assessment research related to such GMO
products. This paper focuses on relevant EPA research and regulatory
examples related to soil effects considerations for GMOs.

Document Number: 350

The chloroplasts of higher plants contain and elaborate many unique
biochemical pathways that produce an astonishing array of compounds that
are vital for plastid function and are also important from agricultural
and nutritional perspectives. One such group of compounds is the
tocochromanols (more commonly known as Vitamin E), which is a class of
four tocopherols and four toctorienols, lipid-soluble antioxidants that
are only synthesized by plants and other oxygenic, photosynthetic
organisms. Though the essential nature of tocopherols in mammalian diets
was recognized over 80 years ago and the biosynthetic pathway in plants
and algae elucidated in the late 1970s and early 80s, it has only been
in the past decade that the genes and proteins for tocopherol synthesis
have finally been isolated and characterized. The use of model plant and
cyanobacterial systems has driven this gene discovery to the point that
manipulation of tocopherol levels and types in various plant tissues and
crops is becoming a reality. This article reviews progress since 1996 in
the molecular and genetic understanding of tocopherol synthesis in the
model photosynthetic organisms Arabidopsis thaliana and Synechocystis
PCC6803 as a primer for current and future efforts to manipulate the
levels of this essential nutrient in food crops by breeding and
transgenic approaches. (c) 2005 Elsevier GmbH. 

Document Number: 351

Bioengineering approaches provide unprecedented opportunities for
reducing poverty, food insecurity, child malnutrition, and natural
resource degradation. Genetic engineering offers outstanding potential
to increase the efficiency of crop improvement. Thus agricultural
biotechnology could enhance global food production and availability in a
sustainable way. Small farmers in developing countries are faced with
many problems and constraints which biotechnology may assist. Yet, there
are varying levels of opposition to the use of this technology in most
countries and it is especially intense in Europe. While there is certain
public apprehension with the use of bioengineering in food improvement,
the primary hurdles facing this technology are the stringent and
burdensome regulatory requirements for commercialization, opposition
from the special interest groups, apprehension by the food industry
especially with the whole foods, and trade barriers including rigid
policies on traceability and labeling. Bioengineered crops such as
soybean, maize, cotton, and canola with a few traits have already made a
remarkable impact on food production and environmental quality. But, in
the developing world, bioengineering of crops such as bananas, cassava,
yams, sweet potatoes, sorghum, rice, maize, wheat, millet, and legumes,
along with livestock, can clearly contribute to global food security.
However, the integration of biotechnology into agricultural research in
developing countries faces many challenges which must be addressed:
financial, technical, political, environmental, activism,
intellectual-property, biosafety, and trade-related issues. To ensure
that developing countries can harness the benefit of this technology
with minimal problems, concerted efforts must be pursued to create an
awareness of its potential benefits and to address the concerns related
to its use through dialog among the various stakeholders: policy makers,
scientists, trade groups, food industry, consumer organizations, farmer
groups, media, and non-governmental organizations. Biotechnology holds
great promise as a new tool in the scientific toolkit for generating
applied agricultural technologies; however, per se it is not a panacea
for the world's problems of hunger and poverty.

Document Number: 352

Pigeonpea [Cajanus cajan (L.) Millsp.] in India had a low compound
growth rate of 0.8% in production between 1949-50 and 2004 due to
various biotic and abiotic stresses. Suitable management technologies to
offset adverse effects of abiotic stresses like waterlogging and drought
have been developed. Soil fertility is another abiotic stress for which
integrated plant nutrient-management system has been evolved to enhance
pigeonpea productivity in pigeonpea-based cropping systems. The
threatening sustainability in cereal-based cropping systems in
North-west Plains Zone, flood menace in North-east Plains Zone and lower
productivity of urdbean or blackgram [Vigna mungo (L.) Hepper] in utera
[sowing in standing rice (Oryza sativa L.) crop] cropping have made way
for pigeonpea-wheat (Triticum aestivum L. emend. Fiori & Paol.), maize
(Zea mays L.)-pre-winter pigeonpea and rice (Oryza sativa L.)-pigeonpea
cropping systems.
The development of about 80 high-yielding, disease-resistant varieties
and hybrids suitable for different situations has helped in improving
not only productivity but also addressing various biotic and abiotic
stresses. Resistant sources have been identified for the diseases like
wilt (Fusarium udum Butler), sterility mosaic virus, blight
(Phytophthora drechsleri Tucker f sp cajani Pal, Grewal & Sarabhoy) and
Alternaria blight [Alternaria alternata (Fr.) Keissler], and pests such
as pod-borer (Helicoverpa armigera. Hubner), pod fly (Melanagromyza
obtusa Malloch) and nematodes, and are being utilized in breeding
programme. The development of pod-borer transgenic by the International
Crops Research Institute for the Semi-Arid Tropics, Patancheru, Andhra
Pradesh, would promote pigeonpea in traditional and non-traditional
areas. Effective cultural and herbicidal methods of weed control to
minimize losses by weeds, and also control methods to minimize losses by
storage pests especially bruchids have also been developed. Processes of
dal making developed at the Central Food Technology Research Institute
and at Pantnagar have shown promise in minimizing the milling losses in
traditional milling.
Effective procurement, attractive minimum support price coupled with
enhanced productivity would promote pigeonpea cultivation in green
revolution tracts. The paper deals with the progress made on various
aspects of pigeonpea production besides highlighting the future research
thrusts.

Document Number: 353

Genetically modified (GM) cotton lines have been approved for
commercialization and widely cultivated in many countries, especially in
China. As a step towards the development of reliable qualitative and
quantitative PCR methods for detecting GM cottons, we report here the
validation of the cotton (Gossypium hirsutum) endogenous reference
control gene, Sad1, using conventional and real-time (RT)-PCR methods.
Both methods were tested on 15 different G. hirsutum cultivars, and
identical amplicons were obtained with all of them. No amplicons were
observed when DNA samples from three species of genus Gossypium,
Arabidopsis thaliana, maize, and soybean and others were used as
amplified templates, demonstrating that these two systems are specific
for the identification and quantification of G. hirsutum. The results of
Southern blot analysis also showed that the Sad1 gene was two copies in
these 15 different G. hirsutum cultivars. Furthermore, one multiplex
RT-quantitative PCR employing this gene as an endogenous reference gene
was designed to quantify the Cry1A(c) gene modified from Bacillus
thuringiensis (Bt) in the insect-resistant cottons, such as Mon531 and
GK19. The quantification detection limit of the Cry1A(c) and Sad1 genes
was as low as 10 pg of genomic DNA. These results indicat that the Sad1
gene can be used as an endogenous reference gene for both qualitative
and quantitative PCR detection of GM cottons.

Document Number: 354

Carotenoids are components of the photosynthetic machinery,
intermediates in the biosynthesis of abscisic acid and other
apocarotenoids and act as coloured pigments particularly in floral and
fruit tissue. Humans benefit in a number of ways from dietary
carotenoids present in green leaf tissue and many fruits, seeds, roots
and tubers. Carotenoids with a beta-ring end group are required for the
synthesis of vitamin A, and deficiency for this vitamin remains a major
health problem in some parts of the world. Epidemiological studies
suggest that carotenoids also have important roles in a range of
diseases including age-related macular degradation and some cancers. The
isoprenoid pathway is described, which leads to the carotenoids via the
condensation of five-carbon isoprenoid units to form a 40-carbon chain
in phytoene. The early part of the biosynthetic pathway involves
crosstalk between plastidic and cytosolic components. Desaturation,
isomerization, cyclization, hydroxylation and epoxidation of the
40-carbon phytoene gives the range of carotenoids found in plants. The
nuclear-encoded enzymes for these stages are targeted to plastids, and
in some cases, different members of a gene family are active in plastids
in different tissue types. Studies on the transgenic manipulation of the
pathway, natural and induced mutants, gene silencing and gene and
quantitative trait locus mapping are increasingly unravelling the
pathway and its control, giving opportunities for directed manipulation
of the types and quantities of these nutritionally important compounds
in crop tissues used in human and animal diets.

Document Number: 355

Two previously developed platforms, a multiplex polymerase chain
reaction (PCR) and a peptide nucleic acid (PNA) array, the former
allowing for the simultaneous detection of five transgenes and two
endogenous controls in food and feed matrices and the latter for the
assessment of the identity of amplified PCR products, were combined in
order to develop a PNA array device for the screening of genetically
modified organisms (GMOs) in food. PNA probes were opportunely designed,
synthesized, and deposited on commercial slides. The length of the
probes as well as the distance of the probes from the surface were
evaluated and found to be critical points. The most suitable probes were
found to be 15-mer PNAs linked to the slide surface by means of two
2-(2-aminoethoxy)-ethoxyacetic acids as spacers. The device was tested
on a model system constituted by flour samples containing a mixture of
standards at known concentrations of transgenic material, in particular
Roundup Ready soybean and Bt11, Bt176, Mon810, and GA21 maize: The DNA
was amplified using the specific multiplex PCR method and tested on the
PNA array. The method proposed was found to be able to correctly
identify every GMO present in the tested samples.

Document Number: 356

Labeling of genetically modified organisms (GMOs) is now in place in
many countries, including the European Union, in order to guarantee the
consumer's choice between GM and non-GM products. Screening of samples
is performed by polymerase chain reaction (PCR) amplification of
regulatory sequences frequently introduced into genetically modified
plants. Primers for the 35S promoter from Cauliflower mosaic virus
(CaMV) are those most frequently used. In virus-infected plants or in
samples contaminated with plant material carrying the virus,
false-positive results can consequently occur. A system for real-time
PCR using a TaqMan minor groove binder probe was designed that allows
recognition of virus coat protein in the sample, thus allowing
differentiation between transgenic and virus-infected samples. We
measured the efficiency of PCR amplification, limits of detection and
quantification, range of linearity, and repeatability of the assay in
order to assess the applicability of the assay for routine analysis. The
specificity of the detection system was tested on various virus isolates
and plant species. All 8 CaMV isolates were successfully amplified using
the designed system. No cross-reactivity was detected with DNA from 3
isolates of the closely related Carnation etched ring virus. Primers do
not amplify plant DNA from available genetically modified maize and
soybean lines or from different species of Brassicaceae or Solanaceae
that are natural hosts for CaMV. We evaluated the assay for different
food matrixes by spiking CaMV DNA into DNA from food samples and have
successfully amplified CaMV from all samples. The assay was tested on
rapeseed samples from routine GMO testing that were positive in the 35S
screening assay, and the presence of the virus was confirmed.

Document Number: 357

Genetically modified (GM) rice (Liberty-Link, event LLRICE62) that is
tolerant to glufosinate ammonium (Liberty) herbicide was compared with a
near-isogenic (NI) conventional medium-grain brown rice (cultivar,
Bengal) and a commercially milled long-grain brown rice in diets for
growing-finishing pigs. The GM and NI rice were grown in 2000. The GM
rice was from fields treated (GM+) or not treated (GM-) with glufosinate
herbicide. The GM- and NI rice were grown using herbicide regimens
typical of southern United States rice production practices. The four
rice grains were similar in composition. Growing-finishing pigs (n = 96)
were fed fortified rice-soybean meal diets containing the four different
rice grains from 25 to 106 kg BW. Diets contained 0.99% lysine initially
(growing phase), with lysine decreased to 0.80% (early finishing phase)
and 0.65% (late finishing phase), when pigs reached 51 and 77 kg,
respectively. The percentage of rice in the four diets was constant
during each of the three phases (72.8, 80.0, and 85.8% for the growing,
early-finishing, and late-finishing phases, respectively). There were
six pen replicates (three pens of barrows and three pens of gilts) and
four pigs per pen for each dietary treatment. All pigs were slaughtered
at the termination of the study to collect carcass data. At the end of
the 98-d experiment, BW gain, feed intake (as-fed basis), and feed:gain
ratio did not differ (P > 0.05) for pigs fed the GM+ vs. conventional
rice diets, but growth performance traits of pigs fed the GM+ rice diets
were superior (P < 0.05) to those of pigs fed the GM- rice diet (ADG =
0.86, 0.79, 0.81, and 0.85 kg/d; ADFI = 2.41, 2.49, 2.37, and 2.45 kg/d;
feed:gain = 2.80, 3.17, 2.95, and 2.89 for GM+, GM-, NI, and
commercially milled rice, respectively). Carcass traits (adjusted for
final BW) did not differ (P = 0.10) among treatments (hot carcass yield
= 73.5, 72.6, 72.6, and 73.2%; 10th-rib backfat = 23.0, 22.7, 21.3, and
23.8 mm; LM area = 38.6, 38.0, 38.2, and 38.1 cm(2); carcass fat-free
lean = 50.5, 50.5, 51.2, and 50.0%). Gilts grew slower (P < 0.05) and
were leaner (P < 0.05) than barrows. Responses to type of rice did not
differ between barrows and gilts, with no evidence of a diet x gender
interaction (P = 0.50) for any trait. The results indicate that the
glufosinate herbicide-tolerant rice was similar in composition and
nutritional value to conventional rice for growing-finishing pigs.

Document Number: 358

Since the 18th of April 2004, two new regulations, EC/1829/2003 on
genetically modified food and feed products and EC/1830/2003 on
traceability and labeling of GMOs, are in force in the EU. This new,
comprehensive regulatory framework emphasizes the need of an adequate
tracing system. Unique identifiers, such as the transgene genome
junction region or a specific rearrangement within the transgene DNA,
should form the basis of such a tracing system. In this study, we
describe the development of event-specific tracing systems for
transgenic maize lines Bt11, Bt176, and GA21 and for canola event GT73.
Molecular characterization of the transgene loci enabled us to clone an
event-specific sequence into a plasmid vector, to be used as a marker,
and to develop line-specific primers. Primer specificity was tested
through qualitative PCRs and dissociation curve analysis in SYBR Green I
real-time PCRs. The primers were then combined with event-specific
TaqMan probes in quantitative real-time PCRs. Calibration curves were
set up both with genomic DNA samples and the newly synthesized plasmid
DNA markers. It is shown that cloned plasmid GMO target sequences are
perfectly suitable as unique identifiers and quantitative calibrators.
Together with an event-specific primer pair and a highly specific TaqMan
probe, the plasmid markers form crucial components of a unique and
straighforward tracing system for Bt11, Bt176, and GA21 maize and GT73
canola events.

Document Number: 359

BACKGROUND AND OBJECTIVE: We have previously demonstrated an antiobesity
effect of dietary Ca; this is largely mediated by Ca suppression of
calcitriol levels, resulting in reduced adipocyte intracellular Ca2+
and, consequently, a coordinated increase in lipid utilization and
decrease in lipogenesis. Notably, dairy Ca is markedly more effective
than other Ca sources.
DESIGN: Obese subjects were placed on balanced deficit (-500 kcal/day)
diets and randomized to control (400-500 mg Ca/day; n = 16) or yogurt
(1100 mg Ca/day; n = 18) treatments for 12 weeks. Dietary macronutrients
and fiber were held constant at the US average.
MEASUREMENTS: Body weight, body fat and fat distribution (by dual-energy
X-ray absorptiometry), blood pressure and circulating lipids were
measured at baseline and after 12 weeks of intervention. RESULTS: Fat
loss was markedly increased on the yogurt diet (-4.43 +/- 0.47 vs -2.75
+/- 0.73 kg in yogurt and control groups; P < 0.005) while lean tissue
loss was reduced by 31% on the yogurt diet. Trunk fat loss was augmented
by 81% on the yogurt vs control diet (P < 0.001), and this was reflected
in a markedly greater reduction in waist circumference (-3.99 +/- 0.48
vs -0.58 +/- 1.04 cm, P < 0.001). Further, the fraction of fat lost from
the trunk was higher on the yogurt diet vs control (P < 0.005).
CONCLUSION: Isocaloric substitution of yogurt for other foods
significantly augments fat loss and reduces central adiposity during
energy restriction.

Document Number: 360

With the development of transgenic crops, regulations to label the
genetically modified organisms (GMOs) and their derived products have
been issued in many countries. Polymerase chain reaction (PCR) methods
are thought to be reliable and useful techniques for qualitative and
quantitative detection of GMOs. These methods are generally needed to
amplify the transgene and compare the amplified results with that of a
corresponding reference gene to get the reliable results. Specific
primers were developed for the rapeseed (Brassica napus),
high-mobility-group protein I/Y(HMG-I/Y) single-copy gene and PCR
cycling conditions suitable for the use of this sequence as an
endogenous reference gene in both qualitative and quantitative PCR
assays. Both methods were assayed with 15 different rapeseed varieties,
and identical amplified products were obtained with all of them. No
amplification was observed when templates were the DNA samples from the
other species of Brassica genus or other species, such as broccoli, stem
mustard, cauliflower, Chinese cabbage, cabbage, sprouts, Arabidopsis
thaliana, carrot, tobacco, soybean, mung bean, tomato, pepper, eggplant,
plum, wheat, maize, barley, rice, lupine, and sunflower. This system was
specific for rapeseed. Limits of detection and quantitation in
qualitative and quantitative PCR systems were about 13 pg DNA (about 10
haploid genomes) and about 1.3 pg DNA (about 1 haploid genome),
respectively. To further test the feasibility of this HMG-I/Y gene as an
endogenous reference gene, samples containing transgenic rapeseed GT73
with the inserted glyphosate oxidoreductase (GOX) gene were quantitated.
These demonstrated that the endogenous PCR detection systems were
applicable to the qualitative and quantitative detection of transgenic
rapeseed.

Document Number: 361

Sweetpotato (Ipomoea batatas) is a widely grown food crop, in which the
most important diseases are caused by viruses. Genetic variability of
three widely distributed sweetpotato viruses was analysed using data
from 46 isolates of Sweet potato feathery mottle virus (SPFMV), 16
isolates of Sweet potato mild mottle virus (SPMMV) and 25 isolates of
Sweet potato chlorotic stunt virus (SPCSV), of which 19, seven and six
isolates, respectively, are newly characterized. Division of SPFMV into
four genetic groups (strains) according to phylogenetic analysis of coat
protein (CP) encoding sequences revealed that strain EA contained the
East African isolates of SPFMV but none from elsewhere. In contrast,
strain RC contained ten isolates from Australia, Africa, Asia and North
America. Strain O contained six heterogeneous isolates from Africa, Asia
and South America. The seven strain C isolates from Australia, Africa,
Asia, and North and South America formed a group that was genetically
distant from the other SPFMV strains. SPMMV isolates showed a high level
of variability with no discrete strain groupings. SPCSV isolates from
East Africa were phylogenetically distant to SPCSV isolates from
elsewhere. Only from East Africa were adequate data available for
different isolates of the three viruses to estimate the genetic
variability of their local populations. The implications of the current
sequence information and the need for more such information from most
sweetpotato-growing regions of the world are discussed in relation to
virus diagnostics and breeding for virus resistance.

Document Number: 362

Transfer of genes from heterologous species provides the means of
selectively introducing new traits into crop plants and expanding the
gene pool beyond what has been available to traditional breeding
systems. With the recent advances in genetic engineering of plants, it
is now feasible to introduce into crop plants, genes that have
previously been inaccessible to the conventional plant breeder, or which
did not exist in the crop of interest. This holds a tremendous potential
for the genetic enhancement of important food crops. However, the
availability of efficient transformation methods to introduce foreign
DNA can be a substantial barrier to the application of recombinant DNA
methods in some crop plants. Despite significant advances over the past
decades, development of efficient transformation methods can take many
years of painstaking research. The major components for the development
of transgenic plants include the development of reliable tissue culture
regeneration systems, preparation of gene constructs and efficient
transformation techniques for the introduction of genes into the crop
plants, recovery and multiplication of transgenic plants, molecular and
genetic characterization of transgenic plants for stable and efficient
gene expression, transfer of genes to elite cultivars by conventional
breeding methods if required, and the evaluation of transgenic plants
for their effectiveness in alleviating the biotic and abiotic stresses
without being an environmental biohazard. Amongst these, protocols for
the introduction of genes, including the efficient regeneration of
shoots in tissue cultures, and transformation methods can be major
bottlenecks to the application of genetic transformation technology.
Some of the key constraints in transformation procedures and possible
solutions for safe development and deployment of transgenic plants for
crop improvement are discussed.

Document Number: 363

Three recent reports (Baoxiu Qi et al., Amine Abbadi et al. and Anthony
J. Kinney et al.) describe the production of very long-chain
polyunsaturated fatty acids in transgenic plants. This might lead to a
sustainable source of these valuable fatty acids for use in human food
and animal feed. At present they are mainly available via consumption of
fish, which is a limited and endangered resource.

Document Number: 364

Flavonoids represent a large and important group of plant natural
products that are ubiquitous in the plant kingdom. Epidemiological
studies have shown the health benefits of a diet high in flavonoids.
However, the dietary intake of flavonoids in most western populations is
limited, creating a need to find alternative food sources for these
polyphenolic secondary metabolites. The domestication of many of our
cultivated food crops has resulted in alterations in the blosynthetic
pathways of many essential micronutrients and vitamins through
inadvertent counterselection against nutritional traits in favor of
agronomic ones. Flavonoids are nearly absent from fruits of cultivated
tomato (Lycopersidon esculentum Mill.), a major vegetable in human
diets. Previous attempts to restore the flavonoid pathway in tomato
fruits have been limited to transgenic strategies, suggesting that the
problem was intractable through traditional methods. Here, we describe
for the first time a nontransgenic metabolic engineering approach to
developing a high flavonoid tomato using a wild tomato species
(Lycopersicon pennellii v. puberulum) and demonstrate the opportunities
for restoring functional pathways using the genetic resources of wild
species, resulting in production of healthier foods.

Document Number: 365

We recently developed a multiplex polymerase chain reaction (PCR) system
for the simultaneous detection of four transgenic maize (MON810, Bt176,
Bt11, and GA21), one transgenic soybean (Roundup Ready), and two control
genes (lectin and zein). Because PCR can lead to ambiguous
interpretations due to low specificity, we have developed the ligation
detection reaction (LDR) combined with a universal array as a molecular
tool to confirm results of PCR analysis. Here, we describe the
PCR-LDR-universal array procedure and demonstrate its specificity in
revealing the presence of transgenic DNA in experimental samples, raw
materials, and commercial foodstuffs.

Document Number: 366

Overexpression of bacterial-derived starch metabolic enzymes in plant
starch storage organs represents a valuable strategy for improving
starch quality, bioprocessing and nutritional value. Transgenic rice
seeds producing a thermostable and bifunctional starch hydrolase,
amylopullulanase (APU) from Thermoanaerobacter ethanolicus 39E, were
generated. Starch in these seeds could be hydrolyzed with optimal
temperatures between 85 and 95degreesC, which resulted in complete
conversion of starch into soluble sugars and production of
protein-enriched flour within a few hours. By expressing various levels
of APU, rice seeds containing reduced amounts of amylose, which is an
important factor affecting starch quality, were obtained without a
significant impact on grain yield. Elevation in granule-bound
pullulanase activity correlates with the reduction of amylose in
developing APU-containing rice seeds. APU was found to be localized
within amyloplasts and in cell walls, which could be the result of
overexpression of APU with a signal peptide. This study establishes
novel approaches to alter starch properties, accelerate bioprocessing of
starch and production of protein-enriched flour from rice seeds, and
could significantly impact the industrial and food uses of cereals.

Document Number: 367

Toward the development of reliable qualitative and quantitative
Polymerase Chain Reaction (PCR) detection methods of transgenic
tomatoes, one tomato (Lycopersicon esculentum) species specific gene,
LAT52, was selected and validated as suitable for using as an endogenous
reference gene in transgenic tomato PCR detection. Both qualitative and
quantitative PCR methods were assayed with 16 different tomato
varieties, and identical amplified products or fluorescent signals were
obtained with all of them. No amplified products and fluorescent signals
were observed when DNA samples from 20 different plants such as soybean,
maize, rapeseed, rice, and Arabidopsis thaliana were used as templates.
These results demonstrated that the amplified LAT52 DNA sequence was
specific for tomato. Furthermore, results of Southern blot showed that
the LAT52 gene was a single-copy gene in the different tested tomato
cultivars. In qualitative and quantitative PCR analysis, the detection
sensitivities were 0.05 and 0.005 ng of tomato genomic DNA,
respectively. In addition, two real-time assays employing this gene as
an endogenous reference gene were established, one for the
quantification of processed food samples derived from nontransgenic
tomatoes that contained degraded target DNA and the other for the
quantification of the junction region of CaMV35s promoter and the
anti-sense ethylene-forming enzyme (EFE) gene in transgenic tomato
Huafan No. 1 samples. All of these results indicated that the LAT52 gene
could be successfully used as a tomato endogenous reference gene in
practical qualitative and quantitative detection of transgenic tomatoes,
even for some processed foods derived from transgenic and nontransgenic
tomatoes.

Document Number: 368

Zinc is a trace element essential for the optimal function of a variety
of biochemical and physiological processes. Its role in healthy aging is
particularly important as it prevents neoplastic cell growth, is
involved in mitotic cell division, DNA and RNA repair. Although zinc is
widely available in food, the daily intake in many persons may be
suboptimal. Other causes of low zinc concentrations may be due to small
bowel conditions that cause mucosal damage and thus decrease absorption.
Chronic diseases associated with alterations in zinc status are
bronchial asthma, rheumatoid arthritis and Alzheimer disease. At present
it is uncertain if therapy with zinc would assist in the management of
these chronic diseases. In view of the important cellular functions of
zinc in the human body, a diet with an adequate zinc content is
beneficial in promoting healthy aging and maintaining good health.

Document Number: 369

Polyphenols in food plants are a versatile group of phytochemicals with
many potentially beneficial activities in terms of disease prevention.
In vitro cell culture experiments have shown that polyphenols possess
antioxidant properties, and it is thought that these activities account
for disease-preventing effects of diets high in polyphenols. However,
polyphenols may be regarded as xenobiotics by animal cells and are to
some extent treated as such, ie, they interact with phase I and phase II
enzyme systems. We recently showed that dietary plant polyphenols,
namely, the flavonoids, modulate expression of an important enzyme in
both cellular antioxidant defenses and detoxification of xenobiotics,
ie, gamma-glutamylcysteine synthetase. This enzyme is rate limiting in
the synthesis of the most important endogenous antioxidant in cells,
glutathione. We showed in vitro that flavonoids increase expression of
-y-glutamylcysteine synthetase and, by using a unique transgenic
reporter mouse strain, we showed increased expression in vivo, with a
concomitant increase in the intracellular glutathione concentrations in
muscles. Because glutathione is important in redox regulation of
transcription factors and enzymes for signal transduction, our results
suggest that polyphenol-mediated regulation of glutathione alters
cellular processes. Evidently, glutathione is important in many
diseases, and regulation of intracellular glutathione concentrations may
be one mechanism by which diet influences disease development. The aim
of this review is to discuss some of the mechanisms involved in the
glutathione-mediated. endogenous, cellular antioxidant defense system,
how its possible modulation by dietary polyphenols such as flavonoids
may influence disease development, and how it can be studied with in
vivo imaging.

Document Number: 370

The major goal of this review is to assess food risks of the
introduction of genetically modified (GM) crops. The author analyzes the
properties of the several classes of target proteins used in the
transgenic constructions and discusses the problems that arise due to
the pleiotropic action of transgenic proteins, the horizontal transfer
of the transgenic constructions, primarily in bacteria, and their
instability. Particular consideration is given to elevated risks of
using the GM plant varieties for producing pharmaceutical preparations,
due to the probability of uncontrolled cross-pollination between the GM
plants and the plants grown for foodstuff production. The author
emphasizes the requirement for assessing in detail all hypothetic risks
in each particular case of cultivating GM varieties; as a control, such
assessment must involve a comprehensive comparison with the conventional
parental forms.

Document Number: 371

Recent developments in mammalian biotechnologies that have been driven
largely by medical bioscience, offer new opportunities for livestock
industries. Major impacts may be expected in the area of reproductive,
genomic and cell technologies that could lead to improved animal
breeding strategies or animal production and health applications. In
particular, the use of advanced reproductive technologies to select
animals at very early stages of life, possibly as early as a 4-day
embryo, combined with genomic technologies to predict genetic merit,
could lead to significantly increased rates of genetic gain. Such
advanced animal breeding technologies will depend strongly on
conventional quantitative genetic evaluation systems. Genetic
modification in the near future will offer targeted animal improvement
options for control of health and production. Long-term impact of
genetic modification on animal production systems will depend on
consumer acceptance, and its perception by social, environmental and
animal welfare groups. However, the opportunity to develop animal
products beyond conventional boundaries may prove too attractive with
genetic modification eventually being accepted as the norm. The
naturally synergistic effect of ex vivo transgenic modification of
embryo stem cell or somatic cell lines, combined with nuclear transfer
present potentially high value propositions for development of novel and
high value products. Opportunities for the mass production of elite
males for use in extensive animal production systems will be possible.

Document Number: 372

A vast number of plant pathogens from viroids of a few hundred
nucleotides to higher plants cause diseases in our crops. Their effects
range from mild symptoms to catastrophes in which large areas planted to
food crops are destroyed. Catastrophic plant disease exacerbates the
current deficit of food supply in which at least 800 million people are
inadequately fed. Plant pathogens are difficult to control because their
populations are variable in time, space, and genotype. Most insidiously,
they evolve, often overcoming the resistance that may have been the
hard-won achievement of the plant breeder. In order to combat the losses
they cause, it is necessary to define the problem and seek remedies. At
the biological level, the requirements are for the speedy and accurate
identification of the causal organism, accurate estimates of the
severity of disease and its effect on yield, and identification of its
virulence mechanisms. Disease may then be minimized by the reduction of
the pathogen's inoculum, inhibition of its virulence mechanisms, and
promotion of genetic diversity in the crop. Conventional plant breeding
for resistance has an important role to play that can now be facilitated
by marker-assisted selection. There is also a role for transgenic
modification with genes that confer resistance. At the political level,
there is a need to acknowledge that plant diseases threaten our food
supplies and to devote adequate resources to their control.

Document Number: 373

The availability of foods low in sugar content yet high in flavour is
critically important to millions of individuals conscious of
carbohydrate intake for diabetic or dietetic purposes. Brazzein is a
sweet protein occurring naturally in a tropical plant that is
impractical to produce economically on a large scale, thus limiting its
availability for food products. We report here the use of a maize
expression system for the production of this naturally sweet protein.
High expression of brazzein was obtained, with accumulation of up to 4%
total soluble protein in maize seed. Purified corn brazzein possessed a
sweetness intensity of up to 1200 times that of sucrose on a per weight
basis. In addition, application tests demonstrated that
brazzein-containing maize germ flour could be used directly in food
applications, providing product sweetness. These results demonstrate
that high-intensity sweet protein engineered into food products can give
sweetener attributes useful in the food industry.

Document Number: 374

Potato (Solanum tuberosum L.) is one of the most important food crops
after wheat (Triticum aestivum :. emend. Fiori & Paol.), maize (Zea mays
L.) and rice (Oryza sativa L.), historically contributing to food and
nutrition security in the World. This tuber crop exists with more than
200 wild species. The cultivated potatoes are tetraploid (2n=4x=48),
display complex tetrasomic inheritance and are propagated asexually
through tubers. Tuberization is a complex physiological process occurred
on underground stolons under the influence of both extrinsic as well as
intrinsic factors. The fascinating reproductive biology characterized by
2n gametes and endosperm balance number concept allows introgression of
traits from wild to cultivated potato species, and the transgenic
approach has further expanded the scope for selective introgression of
genes across the trans-specific and/ or trans-generic barriers. Late
blight [Phytophihora infestans (Mont) de Bary] is the most severe
disease that threats the potato crop worldwide, and next to severity are
the systematic viral diseases, which cause rapid degeneration of seeds.
Therefore, the availability of healthy seeds comprises the most
important aspect for potato production, and recent biotechnological
approaches have been successfully integrated into disease-free
seed-production scheme. In India, more than 80% of potatoes are grown in
sub-tropical plains under conditions distinct from Europe, and this has
only been possible through concerted research at the Central Potato
Research Institute, Shimla, over the past 55 years resulting in the
development of 35 high-yielding potato varieties for diverse
agro-climatic conditions and the innovation of seed plot technique for
augmenting the seed production. India now ranks number three both in
area and production of potato in the world, and is poised to achieve the
maximum annual growth rate in the coming decade. Coincident with neo-
liberalization and rapid urbanization, a growing synergy between private
and public sector investment in processing industry is all set to
sustain and diversify potato production in the country in the 21st
century for securing the food and nutrition.

Document Number: 375

Due to the preparation of new EU legislation on the adventitious and
unavoidable presence of genetically modified seeds in conventional seed
lots, quantification of the contamination of conventional seed lots is
becoming more and more important. Nowadays quantification is performed
by real-time PCR or by statistically based sub-sampling strategies. Due
to the fact that both techniques imply the grinding and homogenisation
of a sample, specific properties and characteristics of seeds that are
important for quantification are lost.
In some cases information is required on the number of transgenic seeds,
the zygosity or the identity of the contaminants. For these purposes we
propose an alternative method for quantification based on a combination
of a three-dimensional pooling of seeds and a qualitative PCR based
screening strategy. The identification of a single transgenic seed among
a population of 1000 requires no more than 30 PCR reactions; one for
each column (10), row (10) and block (10). The window between which
quantification is possible lies between 0% GMO and 0.9% GMO which covers
the labelling thresholds of 0.3%, 0.5% and 0.7% proposed by the EU.

Document Number: 376

Agricultural production almost needs to double in the twenty-first
century, putting tremendous pressure on agricultural resources. Most
food production increases must come from more agricultural
intensification in the South. This advances the need for a new green
revolution: higher productivity and at the same time less pressure on
the environment. Agrobiotechnology can contribute to this double green
revolution. Biotechnology innovations are often scale-neutral and are
therefore suitable for small farmers. Moreover, genetic modification
offers special advantages for crops that have been domesticated for a
very long time and which are therefore quite different from their wild
relatives. However, agrobiotechnology also engenders risks and dangers,
outlined in the paper. Multinational companies show little interest in
small developing countries because the market is small and intellectual
property rights protection is not effective. Not surprisingly, these are
the very countries where food insecurity problems are most acute. In
many developing countries the capacity to conduct biotechnology research
and development is lacking, as is the legal framework for biosafety
testing, patent enforcement and release of transgenic crops. This is
illustrated by a case study on transgenic plantain bananas, developed by
the Catholic University of Leuven, Belgium. The authors argue that legal
and research capacity building are the main priorities. These can be
achieved through public-private and North-South partnerships.

Document Number: 377

Global commercial production of genetically modified (GM) crops has
grown to over 67 million hectares annually, primarily of
herbicide-tolerant and insect protection crop varieties. GM crops are
produced by the insertion of specific genes that either encode a
protein, or a regulatory RNA sequence. A comprehensive safety evaluation
is conducted for each new commercial GM crop, including an assessment of
the potential allergenicity of any newly introduced protein. If the gene
was derived from an allergenic organism, or the protein sequence is
highly similar to a known allergen, immunoassays, e.g., Western blot
assays and enzyme-linked immunosorbent assay tests, are performed to
identify protein-specific IgE binding by sera of individuals allergic to
the gene source, or the source of the sequence-matched allergen.
Although such assays are commonly used to identify previously unknown
allergens, criteria have not been established to demonstrate that a
protein is unlikely to cause allergic reactions. This review discusses
factors that affect the predictive value of these tests, including
clinical selection criteria for serum donors, selection of blocking
reagents to reduce nonspecific antibody binding, inhibition assays to
verify specificity of binding, and scientifically justified limits of
detection (sensitivity) in the absence of information regarding
biological thresholds.

Document Number: 378

A review is presented of 3 murine models and a swine neonatal model used
to investigate immunotherapeutic options. In Model 1, mutation of linear
IgE-binding epitopeS of Ara h 1 for the preparation of a hypoallergenic
Ara h 1 is discussed with respect to expression in transgenic tobacco
plants and correct folding following expression in the pET16b construct.
In Model 2, the mutations of Ara h 1 were assessed for use as an
immunotherapeutic agent. Although some protective benefit was observed
with the modified Ara h 1 protein, animals desensitized with heat-killed
E. coli preparations showed increased protection to challenge. In Model
3, soybean homologs to peanut proteins were investigated to determine if
soybean immunotherapy can potentially provide benefit to peanut-allergic
subjects. Although some protection was provided, additional
experimentation with respect to optimal doses for sensitization and
challenge will need to be investigated. In Model 4, the neonatal swine
model was used to profile different foods (low to moderate to high
sensitizing) similar to food allergies in humans. Evidence suggests such
feasiblity; however, threshold levels for sensitization and allergic
responses will need additional study. In summary, murine and swine
animal models are being used to address immunotherapeutic avenues and
investigation into the mechanisms of food-allergic sensitization.

Document Number: 379

Cassava is the major root crop for a quarter billion subsistence farmers
in sub-Saharan Africa. It is valued for its ability to grow in adverse
environments and the food security it provides. Cassava contains
potentially toxic levels of cyanogenic glycosides (linamarin) which
protect the plant from herbivory and theft. The cyanogens, including
linamarin and its deglycosylated product, acetone cyanohydrin. can be
efficiently removed from the root by various processing procedures.
Short-cuts in processing: which may occur during famines, can result in
only partial removal of cyanogens. Residual cyanogens in cassava foods
may cause neurological disorders or paralysis, particularly in
nutritionally compromised individuals. To address this problem and to
further understand the function of cyanogenic glycosides in cassava. we
have generated transgenic cassava in which cyanogenic glycoside
synthesis has been selectively inhibited in leaves and roots by
antisense expression of CYP79D1/D2 gene fragments. The CYP79D1/D2 genes
encode two highly similar cytochrome P450s that catalyze the
first-dedicated step in cyanogenic glycoside synthesis. Transgenic
plants in which the expression of these genes was selectively inhibited
in leaves had substantially reduced (60-94% reduction) linamarin leaf
levels. Surprisingly, these plants also had a greater than a 99%
reduction in root linamarin content. In contrast. transgenic plants in
which the CYP79D1/D2 transcripts were reduced to non-detectable levels
in roots had normal root linamarin levels. These results demonstrate
that linamarin synthesized in leaves is transported to the roots and
accounts for nearly all of the root linamarin content. Importantly,
transgenic plants having reduced leaf and root linamarin content were
unable to grow in the absence of reduced nitrogen (NH(3)). Cassava roots
have previously been demonstrated to have an active cyanide assimilation
pathway leading to the synthesis of amino acids. We propose that cyanide
derived from linamarin is a major source of reduced nitrogen for cassava
root protein synthesis. Disruption of linamarin transport from leaves in
CYP79D1/D2 anti-sense plants prevents the growth of cassava roots in the
absence of an alternate source of reduced nitrogen. An alternative
strategy for reducing cyanogen toxicity in cassava foods is to
accelerate cyanogenesis and cyanide volatilization during food
processing. To achieve this objective, we have expressed the
leaf-specific enzyme hydroxynitrile lyase (HNL) in roots. HNL catalyzes
the breakdown of acetone cyanohydrin to cyanide. Expression of HNL in
roots accelerated cyanogenesis by more than three-fold substantially
reducing the accumulation of acetone cyanohydrin during processing
relative to wild-type roots.

Document Number: 380

Crop genetic engineering relies on the introduction of foreign DNA into
plant genomes. Although genetically engineered traits provide valuable
alternatives to those available through conventional breeding, there is
public concern about the consumption of foods derived from transgenic
plants. This concern raises the question of whether crops can be
improved by inserting only native DNA into their genomes. Here, we
discuss how rapid advances in molecular biology make it possible to use
plants themselves as DNA sources. Native genes and regulatory elements
can be reintroduced into plants without the need to use selectable
markers. By also using transfer DNAs that are derived from within the
targeted compatibility group, genetically engineered plants can now be
produced that lack any foreign DNA.

Document Number: 381

A nested PCR method has been developed to distinguish traditional
soybean from transgenic Roundup Ready soybeans (RRS). Comparing with the
standard PCR method, the nested PCR method increases the sensitivity by
200 to 1000-fold. One copy of RRS genome can be detected by the nested
PCR. Sufu, a traditional fermented soybean curd, is a very important
Chinese condiment and it is a highly processed food. We have used the
nested PCR to detect seven time points of sufu throughout the aging
period. The nested PCR method showed positive results for sufu made up
of 100% RRS at all seven time points from 0th day to 180th day. This
work describes the development and application of a sensitive and
specific detection method for sufu made from RRS using a nested PCR
system.

Document Number: 382

Transgenic soybean line GTS-40-3-2, marketed under the trade name
Roundup Ready (RR) soy, was developed by Monsanto (USA) to allow for the
use of glyphosate, the active ingredient of the herbicide Roundup, as a
weed control agent. FIR soy was first approved in Canada for
environmental release and for feed products in 1995 and later for food
products in 1996 and is widely grown in Canada. Consumer concern issues
have resulted in proposed labeling regulations in Canada for foods
derived from genetically engineered crops. One requirement for labeling
is the ability to detect and accurately quantify the amount of
transgenic material present in foods. Two assays were evaluated. A
conventional qualitative Polymerase Chain Reaction (PCR) assay to detect
the presence of soy and FIR soy and a real-time PCR to quantify the
amount of RR soy present in samples that tested positive in the first
assay. PCR controls consisted of certified RR soy reference material,
single transgenic soybeans, and a processed food sample containing a
known amount of FIR soy. To test real-world applicability, a number of
common grocery store food items that contain soy-based products were
tested. For some samples, significant differences in amplification
efficiencies during the quantitative PCR assays were observed compared
to the controls, resulting in potentially large errors in
quantification. A correction factor was used to try to compensate for
these differences.

Document Number: 383

Plants can provide most of the nutrients for the human diet. However,
the major crops are often deficient in some of the nutrients. Thus,
malnutrition, with respect to micronutrients such as vitamin A, iron,
and zinc, but also macronutrients such as the essential amino acids
lysine and methionine, affects more than 40% of the world's population.
Recent advances in molecular biology, but also the grasp of biochemical
pathways, metabolic fluxes, and networks can now be exploited to produce
crops enhanced in key nutrients to increase the nutritional value of
plant-derived foods and feeds. Some of the predictions appear to be
accurate, while others not, reflecting the fact that plant metabolism is
more complex than presently understood. A good example for a complex
regulation is the methionine biosynthetic pathway in plants. The
nutritional importance of Met and cysteine has motivated extensive
studies of their roles in plant molecular physiology, especially
regarding to their transport, synthesis, and accumulation in plants.
Recent studies have demonstrated that Met metabolism is regulated
differently in various plant species.

Document Number: 384

Peptide nucleic acid (PNA) microarrays for the detection of Roundup
Ready soybeans in food have been prepared. PNA probes are known to be
more efficient and selective in binding DNA sequences than the analogous
oligonucleotides and are very suitable to be used for diagnostics in
food. PNAs of different lengths were carefully designed and synthesized
by solid-phase synthesis on an automatic synthesizer adopting the BOC
strategy. PNAs were purified by HPLC and characterized by HPLC/ MS. The
probes were spotted on a functionalized surface to produce a microarray
to be hybridized with PCR products. DNA extracted from reference
material was amplified using Cy3- and Cy5-labeled primers, and the
fluorescent PCR products obtained were hybridized on the microarray. Two
protocols were adopted: the hybridization with dsDNA or with ssDNA
obtained by digestion with the enzyme lambda exonuclease. The best
results were obtained using a 15-mer PNA probe in combination with the
ssPCR product derived from enzymatic digestion. The method was applied
to the analysis of a sample of certified transgenic soybean flour.

Document Number: 385

The appearance of genetically modified organisms on the food market a
few years ago, and the demand for more precise and reliable techniques
to detect foreign (transgenic or pathogenic) DNA in edible plants, have
been the driving force for the introduction of real-time PCR techniques
in plant research. This was followed by numerous fundamental research
applications aiming to study the expression profiles of endogenous genes
and multigene families. Since then, the interest in this technique in
the plant scientist community has increased exponentially. This review
describes the technical features of quantitative real-time PCR that are
especially relevant to plant research, and summarizes its present and
future applications.

Document Number: 386

This paper provides guidance on how to assess the safety of foods
derived from genetically modified crops (GM crops); it summarises
conclusions and recommendations of Working Group I of the ENTRANSFOOD
project. The paper provides an approach for adapting the test strategy
to the characteristics of the modified crop and the introduced trait,
and assessing potential unintended effects from the genetic
modification. The proposed approach to safety assessment starts with the
comparison of the new GM crop with a traditional counterpart that is
generally accepted as safe based on a history of human food use (the
concept of substantial equivalence). This case-focused approach ensures
that foods derived from GM crops that have passed this extensive
test-regime are as safe and nutritious as currently consumed
plant-derived foods. The approach is suitable for current and future GM
crops with more complex modifications. First, the paper reviews test
methods developed for the risk assessment of chemicals, including food
additives and pesticides, discussing which of these methods are suitable
for the assessment of recombinant proteins and whole foods. Second, the
paper presents a systematic approach to combine test methods for the
safety assessment of foods derived from a specific GM crop. Third, the
paper provides an overview on developments in this area that may prove
of use in the safety assess-merit of GM crops, and recommendations for
research priorities. It is concluded that the combination of existing
test methods provides a sound test-regime to assess the safety of GM
crops. Advances in our understanding of molecular biology, biochemistry,
and nutrition may in future allow further improvement of test methods
that will over time render the safety assessment of foods even more
effective and informative. (C) 2004 Elsevier Ltd. 

Document Number: 387

In 2000, the thematic network ENTRANSFOOD was launched to assess four
different topics that are all related to the testing or assessment of
food containing or produced from genetically modified organisms (GMOs).
Each of the topics was linked to a European Commission (EC)-funded large
shared cost action (see http://www.entransfood.com). Since the exchange
of genetic information through horizontal (lateral) gene transfer (HGT)
might play a more important role, in quantity and quality, than hitherto
imagined, a working group dealing with HGT in the context of food and
feed safety was established. This working group was linked to the
GMOBILITY project (GMOBILITY, 2003) and the results of the deliberations
are laid down in this review paper. HGT is reviewed in relation to the
potential risks of consuming food or feed derived from transgenic crops.
First, the mechanisms for obtaining transgenic crops are described.
Next, HGT mechanisms and its possible evolutionary role are described.
The use of marker genes is presented in detail as a special case for
genes that may pose a risk. Furthermore, the exposure to GMOs and in
particular to genetically modified (GM) deoxyribonucleic acid (DNA) is
discussed as part of the total risk assessment. The review finishes off
with a number of conclusions related to GM food and feed safety. The aim
of this paper is to provide a comprehensive overview to assist risk
assessors as well as regulators and the general public in understanding
the safety issues related to these mechanisms. (C) 2004 Elsevier Ltd.

Document Number: 388

Both labelling and traceability of genetically modified organisms are
current issues that are considered in trade and regulation. Currently,
labelling of genetically modified foods containing detectable transgenic
material is required by EU legislation. A proposed package of
legislation would extend this labelling to foods without any traces of
transgenics. These new legislations would also impose labelling and a
traceability system based on documentation throughout the food and feed
manufacture system. The regulatory issues of risk analysis and labelling
are currently harmonised by Codex Alimentarius. The implementation and
maintenance of the regulations necessitates sampling protocols and
analytical methodologies that allow for accurate determination of the
content of genetically modified organisms within a food and feed sample.
Current methodologies for the analysis of genetically modified organisms
are focused on either one of two targets, the transgenic DNA inserted-
or the novel protein(s) expressed- in a genetical]v modified product.
For most DNA-based detection methods, the polymerase chain reaction is
employed. Items that need consideration in the use of DNA-based
detection methods include the specificity, sensitivity, matrix effects,
internal reference DNA, availability of external reference materials,
hemizygosity versus homozygosity, extrachromosomal DNA, and
international harmonisation. For most protein-based methods,
enzyme-linked immunosorbent assays with antibodies binding the novel
protein are employed. Consideration should be given to the selection of
the antigen bound by the antibody, accuracy, validation, and matrix
effects. Currently, validation of detection methods for analysis of
genetically modified organisms is taking place. In addition, new
methodologies are developed, including the use of microarrays, mass
spectrometry, and surface plasmon resonance. Challenges for GMO
detection include the detection of transgenic material in materials with
varying chromosome numbers. The existing and proposed regulatory EU
requirements for traceability of genetically modified products fit
within a broader tendency towards traceability of foods in general and,
commercially, towards products that can be distinguished from each
other. Traceability systems document the history of a product and may
serve the purpose of both marketing and health protection. In this
framework, segregation and identity preservation systems allow for the
separation of genetically modified and non-modified products from "farm
to fork". Implementation of these systems comes with specific technical
requirements for each particular step of the food processing chain. In
addition, the feasibility of traceability systems depends on a number of
factors, including unique identifiers for each genetically modified
product, detection methods, permissible levels of contamination, and
financial costs. In conclusion, progress has been achieved in the field
of sampling, detection, and traceability of genetically modified
products, while some issues remain to be solved. For success, much will
depend on the threshold level for adventitious contamination set by
legislation. (C) 2004 Published by Elsevier Ltd.

Document Number: 389

The most important results from the EU-sponsored ENTRANSFOOD Thematic
Network project are reviewed, including the design of a detailed
step-wise procedure for the risk assessment of foods derived from
genetically modified crops based on the latest scientific developments,
evaluation of topical risk assessment issues, and the formulation of
proposals for improved risk management and public involvement in the
risk analysis process. (C) 2004 Elsevier Ltd. 

Document Number: 390

The detection of genetically modified organisms (GMOs) in food and feed
is an important issue for all the subjects involved in raw material
control, food industry, and distribution. Because the number of GMOs
authorized in the EU increased during the past few years, there is a
need for methods that allow a rapid screening of products. In this
paper, we propose a method for the simultaneous detection of four
transgenic maize (MON810, Bt11, Bt 176, and GA21) and one transgenic
soybean (Roundup Ready), which allows routine control analyses to be
sped up. DNA was extracted either from maize and soybean seeds and
leaves or reference materials, and the recombinant DNA target sequences
were detected with 7 primer pairs, accurately designed to be highly
specific for each investigated transgene. Cross and negative controls
were performed to ensure the specificity of each primer pair. The method
was validated on an interlaboratory ring test and good analytical
parameters were obtained (LOD = 0.25%, Repeatability, (r) = 1;
Reproducibility, (R) = 0.9). The method was then applied to a model
biscuit made of transgenic materials baked for the purpose and to real
samples such as feed and foodstuffs. On account of the high recognition
specificity and the good detection limits, this multiplex PCR represents
a fast and reliable screening method directly applicable in all the
laboratories involved in raw material and food control.

Document Number: 391

With the development of transgenic crops, many countries have issued
regulations to label the genetically modified organisms (GMOs) and their
derived products. Polymerase Chain Reaction (PCR) methods are thought to
be reliable and useful techniques for qualitative and quantitative
detection of GMOs. These methods generally need to amplify the transgene
and compare the amplified result with that of the corresponding
reference gene to obtain reliable results. In this article, we reported
the development of specific primers and probe for the rice (Oryza
sativa) sucrose phosphate synthase (SPS) gene and PCR cycling conditions
suitable for the use of this sequence as an endogenous reference gene in
both qualitative and quantitative PCR assays. Both methods were assayed
with 13 different rice varieties, and identical amplification products
were obtained with all of them. No amplification products were observed
when DNA samples from other species, such as wheat, maize, barley,
tobacco, soybean, rapeseed, tomato, sunflower, carrot, pepper, eggplant,
lupine, mung bean, plum, and Arabidopsis thaliana, were used as
templates, which demonstrated that this system was specific for rice. In
addition, the results of the Southern blot analysis confirmed that the
SPS gene was a single copy in the tested rice varieties. In qualitative
and quantitative PCR analyses, the detection sensitivities were 0.05 and
0.005 ng of rice genomic DNA, respectively. To test the practical use of
this SPS gene as an endogenous reference gene, we have also quantified
the beta-glucuronidase (GUS) gene in transgenic rice using this
reference gene. These results indicated that the SPS gene was species
specific, had one copy number, and had a low heterogeneity among the
tested cultivars. Therefore, this gene could be used as an endogenous
reference gene of rice and the optimized PCR systems could be used for
practical qualitative and quantitative detection of transgenic rice.

Document Number: 392

Two separate multiplex PCR assays were developed for the detection of
transgenic papaya and transgenic squash. Papaya line 55-1 contains a
genetic insertion consisting of the coat protein gene of Papaya ringspot
virus strain p and is resistant to infection by this virus. A multiplex
PCR was developed to specifically amplify the papaya ringspot virus coat
protein transgene construct and an endogenous papaya gene sequence. A
third primer set was designed to amplify the beta-glucuronidase gene
construct, which can distinguish between the commercial and
noncommercial papaya lines 55-1 and 63-1. Squash line ZW-20 contains
genetic insertions of the coat protein genes from Watermelon mosaic
virus II and Zucchini yellow mosaic virus and is resistant to these two
viruses. Squash line CZW-3 is similar to ZW-20 but additionally confers
resistance to Cucumber mosaic virus. A second multiplex assay was
developed to detect and differentiate squash lines ZW-20 and CZW-3 in a
single reaction.

Document Number: 393

Food insecurity is one of the most important social issues faced today,
with 840 million individuals enduring chronic hunger and three billion
individuals suffering from nutrient deficiencies. Most of these
individuals are poverty stricken and live in developing countries.
Strategies to address food insecurity must aim to increase agricultural
productivity in the developing world in order to tackle poverty, and
must provide long-term improvements in crop yields to keep up with
demand as the world's population grows. Genetically enhanced plants
provide one route to sustainable higher yields, either by increasing the
intrinsic yield capability of crop plants or by protecting them from
biotic and abiotic constraints. The present paper discusses a range of
transgenic approaches that could increase agricultural productivity if
applied on a large scale, including the introduction of genes that
confer resistance to pests and diseases, or tolerance of harsh
environments, and genes that help to lift the intrinsic yield capacity
by increasing metabolic flux towards storage carbohydrates, proteins and
oils. The paper also explores how the nutritional value of plants can be
improved by genetic engineering. Transgenic plants, as a component of
integrated strategies to relieve poverty and deliver sustainable
agriculture to subsistence farmers in developing countries, could have a
significant impact on food security now and in the future.

Document Number: 394

Polymerase chain reaction (PCR) screening for presence of transgenic
components in food is becoming a routine method in modern food analysis.
To develop a high throughput method for quantitation of the PCR products
is needed for automatic industry analysis. Here we described an in situ
liquid-phase hybridization (LPH) for PCR-enzyme linked immunoabsorbent
assays (PCR-ELISA) that was widely used for quantitation of PCR
products. In LPH-PCR-ELISA, the biotinylated PCR product was hybridized
with digoxigenin-labeled probes in the PCR reaction mixture immediately
after PCR cycles and the hybridizations was incorporated into the PCR
program. Subsequent enzyme conversion of substrate gave distinct OD
values when detecting samples with genetically modified organisms (GMOs)
labels in the different concentrations. The described method enabled a
fast, specific, and accurate detection of GMOs components in food
products and thus can be developed to a full-automatic method for
routine analysis of raw and processed food products in large sample
number. (C) 2003 Elsevier Ltd. 

Document Number: 395

Possible horizontal gene transfer between genetically modified (GM)
plant and food, and various microorganisms including Escherichia coli,
Enterococcus faecalis, and Lactobacilli was investigated. When plasmids
harboring antibiotic-resistant gene were incubated with microorganisms,
no antibiotic-resistant colony appeared on solid media containing
antibiotics (ampicillin and kanamycin), an indication that gene transfer
event from plasmids to microorganisms by natural transformation is rare.
Korean traditional fermented food, Deonjang, was made from GM soybean
using Aspergillus oryzae and Bacillus subtilis. During aging periods,
microorganisms were collected from Deonjang, and
5-enol-pyruvylshikimate-3-phosphate synthase (CP4EPSPS) gene was
detected by PCR. Absence of CP4EPSPS gene in purified microbial DNA was
confirmed by PCR. Horizontal gene transfer event from genetically
modified food to microorganisms was not occurred by natural
transformation in this case study.

Document Number: 396

Maize (Zea mays L.) plants have been transformed to express a Cry1F
insecticidal crystal protein originally isolated from Bacillus
thuringiensis Berliner. This protein controls lepidopteran pests of
maize, including the European corn borer, Ostrinia nubilalis (Hubner).
As part of the safety assessment for crops containing transgenes, a
compositional analysis of the food and feed is conducted. This analysis
is designed to detect unintended changes in the nutrient and
antinutrient content of the raw commodities produced by the crop due to
the insertion of the genes into the genomic DNA of the plant (pleotropic
effects). Samples of transgenic and nontransgenic maize forage and grain
were collected from six field sites located in the U.S. and Canada.
Forage samples were analyzed for proximates and minerals, and grain was
further analyzed for fatty acids, amino acids, vitamins, secondary
metabolites, and antinutrients. Results demonstrated that maize
expressing the Cry1F protein was equivalent to nontransgenic maize with
respect to these important components. Comparison of the variability
within the nontransgenic and transgenic hybrid, as compared to
composition values reported in the literature, suggest that factors
other than transgenes may contribute more substantially to the
composition of crops.

Document Number: 397

The use of transgenic crops as feeds for ruminant animals has prompted
study of the possible uptake of transgene fragments by ruminal
micro-organisms and/or intestinal absorption of fragments surviving
passage through the rumen. The persistence in buffered ruminal contents
of seven different recombinant DNA fragments from GM rapeseed expressing
the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) transgene was
tracked using PCR. Parental and transgenic (i.e. glyphosphate-tolerant;
Roundup Ready(R), Monsanto Company, St Louis, MO, USA) rapeseed were
incubated for 0, 2, 4, 8, 12, 24 and 48 h as whole seeds, cracked seeds,
rapeseed meal, and as pelleted, barley-based diets containing 65 g
rapeseed meal/kg. The seven transgene fragments ranged from 179 to 527
bp and spanned the entire 1363 bp EPSPS transgene. A 180 bp
ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) small subunit
fragment and a 466 bp 16S rDNA fragment were used as controls for
endogenous rapeseed DNA and bacterial DNA respectively. The limit of
detection of the PCR assay, established using negative controls spiked
with known quantities of DNA, was 12.5 pg. Production of gas and NH3 was
monitored throughout the incubation and confirmed active in vitro
fermentation. Bacterial DNA was detected in all sample types at all time
points. Persistence patterns of endogenous (Rubisco) and recombinant
(EPSPS) rapeseed DNA were inversely related to substrate digestibility
(amplifiable for 48, 8 and 4 h in whole or cracked seeds, meal and diets
respectively), but did not differ between parental and GM rapeseed, nor
among fragments. Detection of fragments was representative of
persistence of the whole transgene. No EPSPS fragments were amplifiable
in microbial DNA, suggesting that transformation had not occurred during
the 48 h incubation. Uptake of transgenic DNA fragments by ruminal
bacteria is probably precluded or time-limited by rapid degradation of
plant DNA upon plant cell lysis.

Document Number: 398

Alkaline-cooked corn, called nixtamal, is the basis for many traditional
corn products such as tortillas, chips, and taco shells that are used
widely in Mexico and Central America and in the preparation of snack
foods that are consumed globally. To assess the effects of alkaline and
thermal treatments on the detectability of DNA and protein for the
presence of genetically modified sequences, various nixtamalized
products were prepared from. blends of conventional white corn
containing 0.1, 1.0, and 10% transgenic corn (event CBH 351,
StarLink(TM)). Real-time quantitative polymerase chain reactions
(RTQ-PCR) and immunoassays were used to determine the cry9C gene and
protein, respectively, in unprocessed corn kernels, freshly prepared,
alkaline-cooked and ground corn (mass), masa flour, tortillas prepared
from masa by heat treatment, chips prepared from damp masa dough by:
deep frying, and from tortillas processed at high (200degreesC) and low
temperatures (70degreesC). In spite of progressive degradation of
genomic DNA during processing, RTQ-PCR genetic analysis. allowed
detection and quantification of the cry9C,gene in all products prepared
from 10, 1, and 0.1% StarLink corn, except deep-fried chips containing
0.1% StarLink. Enzyme-linked immunosorbent assays readily detected <1
ppm cry9C protein in all blends of unprocessed corn (10, 1, and 0.1%
StarLink) as well as in nonfried tortilla and masa products. This
technique was not suitable for thermally treated nixtamalized products
containing <1% transgenic corn.

Document Number: 399

An ambitious aim in plant breeding and biotechnology is to increase the
protein content of crop seeds used for food and feed. Using an approach
to manipulate assimilate partitioning, we succeeded in elevating the
protein content in legume seeds up to 50%. Transgenic bean plants were
generated which express a Corynebacterium glutamicum phosphoenolpyruvate
carboxylase (PEPC) in a seed-specific manner. The bacterial enzyme was
not feedback inhibited by malate. Transgenic seeds showed a higher
[C-14]-CO2 uptake and about a threefold increased incorporation of
labelled carbon into proteins. Changed metabolite profiles of maturing
cotyledons indicated a shift of metabolic fluxes from sugars/starch into
organic acids and free amino acids. These changes were consistent with
an increased carbon flow through the anaplerotic pathway catalysed by
PEPC. Consequently, transgenic seeds accumulated up to 20% more protein
per gram seed dry weight. Additionally, seed dry weight was higher by
20%-30%. We conclude that PEPC in seeds is a promising target for
molecular plant breeding.

Document Number: 400

Plant genetic engineering has, until now, relied on the incorporation of
foreign DNA into plant genomes. Public concern about the extent to which
transgenic crops differ from their traditionally bred counterparts has
resulted in molecular strategies and gene choices that limit, but not
eliminate, the introduction of foreign DNA. Here, we demonstrate that a
plant-derived (P-) DNA fragment can be used to replace the universally
employed Agrobacterium transfer (T-) DNA. Marker-free P-DNAs are
transferred to plant cell nuclei together with conventional T-DNAs
carrying a selectable marker gene. By subsequently linking a positive
selection for temporary marker gene expression to a negative selection
against marker gene integration, 29% of derived regeneration events
contain P-DNA insertions but lack any copies of the T-DNA. Further
refinements are accomplished by employing Q-mutated virD2 and
isopentenyl transferase cytokinin genes to impair T-DNA integration and
select against backbone integration, respectively. The presented methods
are used to produce hundreds of marker-free and backbone-free potato
(Solanum tuberosum) plants displaying reduced expression of a
tuber-specific polyphenol oxidase gene in potato. The modified plants
represent the first example of genetically engineered plants that only
contain native DNA.

Document Number: 401

Availability of water is the most limiting factor for crop productivity
and yield. Therefore, there is a strong need to understand plant
adaptation mechanisms against adverse environmental conditions to
improve stress tolerance. Plant stress responses involve the expression
of a plethora of genes with an adaptive role. Their encoded products
comprise enzymes catalyzing the synthesis of osmoprotectants or
antioxidants, lipid desaturases, water channels, and ion transporters
among others. Here we show the characterization of 2 different genes.
First, the CpMYB10 transcription factor gene from the resurrection plant
Craterostigma plantagineum is rapidly induced by dehydration and ABA
treatments in leaves and roots. Transgenic Arabidopsis thaliana plants,
transformed with CpMYB10 promoter fused to GUS gene, showed reporter
expression under ABA and stress conditions in several organs.
Overexpression of CpMYB10 cDNA in Arabidopsis led to desiccation and
salt tolerance of transgenic lines, which also exhibited
glucose-insensitive and ABA-hypersensitive phenotypes. Also, the
trehalose-6-phosphate synthase (AtTPS1) gene overexpressed in
Arabidopsis conferred dehydration tolerance, and glucose- and
ABA-insensitive phenotypes. Transgenic seedlings germinated on high
concentrations of glucose developed normally, in contrast to wild type
seedlings. Germination in the presence of ABA revealed higher
germination rate for transgenic plants overexpressing AtTPS1.
Gene-expression analysis in transgenic plants overexpressing AtTPS1
showed up- or down-regulation of several genes involved in glucose
sensing and ABA signaling. Therefore, our results provide evidence that
CpMYB10 and AtTPS1, when overexpressed in Arabidopsis, led to a
stress-tolerance phenotype through a cross-talk to ABA and glucose
signaling pathways.

Document Number: 402

The ability to express foreign genes using transgenic technologies has
opened up options for producing large quantities of commercially
important industrial or pharmaceutical products in plants. These
technologies have made it possible to use well-developed systems of
commercial agriculture that were developed principally to produce raw
material for large-scale food, feed or processing applications for the
production of foreign molecules. The possibility of the novel industrial
or pharmaceutical molecules produced in such plants, or components
derived from them, contaminating the environment and food chains has
become especially controversial. This potential contamination has
prompted detailed consideration of how such crops and the molecules that
they produce can be effectively isolated and contained. First, the crop
can be completely isolated physically from its food or feed counterpart
during every aspect of its development and commercialization. Second,
genetic isolation systems or genetic barriers that prevent normal
reproduction can be used to reduce the likelihood of the industrial or
pharmaceutical crop entering the food chain.

Document Number: 403

Starches derived from five genetically modified potato lines, two
chemically modified potato starches and two native starches from potato
and maize were subjected to physical and chemical analyses and their
functionality evaluated in a milk-based food product model. The
transgenic starches were specifically modified with respect to
amylopectin chain length and phosphorous content by suppression of the
starch branching enzyme and overexpression of glycogen branching enzyme.
Transgenic starches with long amylopectin chains and high phosphorous
content had increased gelatinisation temperatures, produced gels with a
higher tendency to retrograde and a low freeze/thaw stability as
compared to starches with shorter amylopectin chains and lower
phosphorous content. The textural properties of the food product model
prepared from genetically and chemically modified starches were
characterised by sensory and rheological analyses. To clearly visualise
the effects of the modifications, data was evaluated by radar plots and
multiple regression analysis (chemometrics). Genetically modified potato
starches with longer amylopectin chains and increased phosphorous
content gave a more gelled and a shorter texture as compared to starches
with shorter amylopectin chains and decreased phosphorous content.
Acetylated and hydroxypropylated potato starches gave sticky and stringy
textures. Correlations between rheology parameters and sensory
parameters were found. The sensory parameter stringy/long could be
predicted from the rheological data.

Document Number: 404

The global demand for food is increasing because of the growing world
population. At the same time, availability of arable land is shrinking.
Traditional plant breeding methods have made and will continue to make
important contributions toward meeting the need for more food. In many
areas of the world, however, the problem is food quality. There may be
enough energy available from food, but the staple foods lack certain
essential nutrients. In the developed world, demand for "functional
foods" ( that is, foods that provide health benefits beyond basic
nutrition) is increasing. Nutritional improvements in foods could help
to meet both of these demands for improved food quality. Modern
agricultural biotechnology, which involves the application of cellular
and molecular techniques to transfer DNA that encodes a desired trait to
food and feed crops, is proving to be a powerful complement to
traditional methods to meet global food requirements. An important
aspect of biotechnology is that it provides access to a broad array of
traits that can help meet this need for nutritionally improved
cultivars. The new varieties developed through modern biotechnology have
been identified by a number of terms, including genetically modified (GM
or GMO), genetically engineered (GE or GEO), transgenic, biotech,
recombinant, and plants with novel traits (PNTs). For the present
discussion, the term "GM" will be used because of its simplicity and
broad public recognition.

Document Number: 405

We have applied the ligation detection reaction (LDR) combined with a
universal array approach to the detection and quantitation of the
polymerase chain reaction (PCR) amplified cry1A(b) gene from Bt-176
transgenic maize. We demonstrated excellent specificity and high
sensitivity. Down to 0.5 fmol (nearly 60 pg) of PCR amplified transgenic
material was inequivocally detected with excellent linearity within the
0.1-2.0% range with respect to wild-type maize. We suggest the
feasibility of extending the LDR/universal array format to detect in
parallel several transgenic sequences that are being developed for food
applications.

Document Number: 406

Plant proteins, when used as dietary protein, are generally incomplete
in nutrition due to their deficiency in several essential amino acids,
for example, lysine and tryptophan in cereals and methionine and
cysteine in legumes. Attempts to breed crops with increased levels of
lysine and methionine have been less than satisfactory. Modem
biotechnology offers alternative approaches for rectifying this
nutrition deficiency. In the past decade, several transgenic strategies
aimed at modifying the amino acid composition of plant proteins and
enhancing the content of specific essential amino acid(s) for nutrition
improvement have been developed and tested. These include synthetic
proteins, modification of protein sequences, over-expression of
heterologous or homologous proteins, and metabolic engineering of the
free essential amino acid pool and protein sink. The progress and
potential of these approaches and studies are reviewed. As plant
proteins are the primary source of all dietary protein consumed by
humans and animals and are inexpensive to produce in comparison with
meat, improving their quality will make a significant contribution to
our future food needs. The research and development in this area of
interest is making promising progress towards this endeavor.

Document Number: 407

The global demand for food is increasing because of the growing world
population. At the same time, availability of arable land is shrinking.
Traditional plant breeding methods have made and will continue to make
important contributions toward meeting the need for more food. In many
areas of the world, however, the problem is food quality. There may be
enough energy available from food, but the staple! foods lack certain
essential nutrients. In the developed world, demand for "functional
foods" (that is, foods that provide health benefits beyond basic
nutrition) is increasing. Nutritional improvements in foods could help
to meet both of these demands for improved food quality. Modern
agricultural biotechnology, which involves the application of cellular
and molecular techniques to transfer DNA that encodes a desired trait to
food and feed crops, is proving to be a powerful complement to
traditional methods to meet global food requirements. An important:
aspect of biotechnology is that it provides access to a broad array of
traits that can help meet this need for nutritionally improved
cultivars. The new varieties developed through modern biotechnology have
been identified by a number of terms, including genetically modified (GM
or GMO), genetically engineered (GE or GEO), transgenic, biotech,
recombinant, and plants with novel traits (PNTs). For the present
discussion, the term "GM" will be used because of its simplicity and
broad public recognition.

Document Number: 408

Over three billion people are currently micronutrient (i.e.
micronutrient elements and vitamins) malnourished, resulting in
egregious societal costs including learning disabilities among children,
increased morbidity and mortality rates, lower worker productivity, and
high healthcare costs, all factors diminishing human potential,
felicity, and national economic development. Nutritional deficiencies
(e.g. iron, zinc, vitamin A) account for almost two-thirds of the
childhood death worldwide. Most of those afflicted are dependent on
staple crops for their sustenance. Importantly, these crops can be
enriched (i.e. 'biofortified') with micronutrients using plant breeding
and/or transgenic strategies, because micronutrient enrichment traits
exist within their genomes that can to used for substantially increasing
micronutrient levels in these foods without negatively impacting crop
productivity. Furthermore, 'proof of concept' studies have been
published using transgenic approaches to biofortify staple crops (e.g.
high beta-carotene 'golden rice' grain, high ferritin-Fe rice grain,
etc). In addition, micronutrient element enrichment of seeds can
increase crop yields when sowed to micronutrient-poor soils, assuring
their adoption by farmers. Bioavailability issues must be addressed when
employing plant breeding and/or transgenic approaches to reduce
micronutrient malnutrition. Enhancing substances (e.g. ascorbic acid,
S-containing amino acids, etc) that promote micronutrient
bioavailability or decreasing antinutrient substances (e.g. phytate,
polyphenolics, etc) that inhibit micronutrient bioavailability, are both
options that could be pursued, but the latter approach should be used
with caution. The world's agricultural community should adopt plant
breeding and other genetic technologies to improve human health, and the
world's nutrition and health communities should support these efforts.
Sustainable solutions to this enormous global problem of 'hidden hunger'
will not come without employing agricultural approaches.

Document Number: 409

This article examines the opportunities for using several forms of
modern biotechnology to improve orphan crops in developing countries.
These crops, including tef, millets, cowpea, and indigenous vegetables,
fruits, roots, and tubers, tend to be locally important, but receive
little public or private investment. Recent advances in the fields of
genetics and genomics provide a more unified understanding of the
biology of plants. We summarize some important ways in which genetic
technologies can be harnessed for orphan crops and provide examples of
potential genetic and genomics research that is likely to benefit poor
regions. Finally, we suggest policies that could help create incentives
for application of advanced science to orphan crops. (C) 2004 Elsevier
Ltd. 

Document Number: 410

Over the past 20 years we have generated stable lines of transgenic
Atlantic salmon possessing either antifreeze protein (AFP) genes or a
salmon growth hormone (GH) gene construct. The AFP gene transfer studies
were initiated in 1982. The AFP transgene integrated into salmon genomic
DNA and AFP has been found in the blood of all 5 generations to date.
However, AFP levels are low and a means to raise these levels needs to
be developed. Our GH gene transfer studies were initiated in 1989.
Evidence to date indicates that a single copy of the GH transgene
integrated into chromosomal DNA and has been passed down in Mendelian
fashion, along with its rapid growth phenotype, over 6 generations.
Laboratory studies indicate that our GH transgene enhances growth rates
with Atlantic salmon reaching market size ( 4 - 6 kg) a year earlier
than non-transgenics cultured commercially in Atlantic Canada.
This GH gene transfer technology was patented and licensed to Aqua
Bounty Farms Inc., and the transgenic salmon are currently under review
by various government regulatory authorities in the USA and Canada for
use in commercial aquaculture ventures. Our experience with the
regulatory authorities, the industry and the press indicates that the
successful introduction of transgenic salmon into the aquaculture
industry involves issues concerning not only science but also food
safety, environmental safety, animal welfare and consumer acceptance.
This communication centres on our experience with Atlantic salmon and
outlines our plans and progress towards demonstrating the safety of
transgenic fish to the consumer and to the environment.

Document Number: 411

In this article, the different DNA amplification techniques that are
being used for detecting genetically modified organisms (GMOs) in foods
are examined. This study intends to provide an updated overview
(including works published till June 2002) on the principal applications
of such techniques together with their main advantages and drawbacks in
GMO detection in foods. Some relevant facts on sampling, DNA isolation,
and DNA amplification methods are discussed. Moreover these analytical
protocols are discussed from a quantitative point of view, including the
newest investigations on multiplex detection of GMOs in foods and
validation of methods.

Document Number: 412

Maize GA21 line has integrated several tandemly repeated copies of the
r-act 5-enol-pyruvylshikimate-3-phosphate synthase construct used for
plant transformation. We were able to amplify a nucleotide sequence
corresponding to the polylinker plasmid vector flanked by the r-act
promoter and nopaline synthase 3'-terminator. A method for specific
detection and quantification of Roundup Ready(R) transgenic maize line
GA21 DNA using conventional and real-time PCR and based on this
transgenic sequence is described. GA21 specific primers and probe were
designed targeting the vector-promoter junction region and amplifying a
72-bp DNA fragment. Quantification methods were optimized through three
different real-time PCR chemistries, i.e. SYBR(R) Green I,
Amplifluor(TM) and TaqMan(R). All three methods proved to be specific,
highly sensitive and reliable for both identification and quantification
of GA21 DNA.
Plasmid pGAivr containing single copies of the GA21 and invertase
amplicons was constructed for use as external standard in calibration
curves. Using pGAivr, a TaqMan(R) based real-time PCR assay was
optimized in duplex format targeting the maize species-specific ivr1
gene and the GA21 junction region. The detection limit of the method was
0.01% GA21, which is far below the established threshold for accidental
presence of genetically modified organisms (GMO), this method therefore
being suitable for use in routine GMO analysis. (C) 2003 Elsevier Ltd.

Document Number: 413

Wheat, barley, rye free diet is required to control coeliac disease and
therefore adequate analytical tools are necessary to check gluten-free
products. At the moment, official analytical methods are based on
immunochemical assays for gliadin detection. However, DNA-based methods
for detection of specific sequences have been extensively proposed as
alternative to the protein-based ones, e.g. for transgenic contamination
analysis. The aim of this work has been to develop an alternative
quantification method for rye-content evaluation in raw materials and
processed food based on real-time PCR approach using both SYBR green
detection system and TaqMan fluorogenic probe. (C) 2003 Swiss Society of
Food Science and Technology. Published by Elsevier Ltd. All rights
reserved.

Document Number: 414

Transgenic resistance to insects has been demonstrated in plants
expressing insecticidal genes such as delta-endotoxins from Bacillus
thuringiensis (Bt), protease inhibitors, enzymes, secondary plant
metabolites, and plant lectins. While transgenic plants with introduced
Bt genes have been deployed in several crops on a global scale, the
alternative genes have received considerably less attention. The
protease inhibitor and lectin genes largely affect insect growth and
development and, in most instances, do not result in insect mortality.
The effective concentrations of these proteins are much greater than the
Bt toxin proteins. Therefore, the potential of some of the alternative
genes can only be realized by deploying them in combination with
conventional host plant resistance and Bt genes. Genes conferring
resistance to insects can also be deployed as multilines or synthetic
varieties. Initial indications from deployment of transgenics with
insect resistance in diverse cropping systems in USA, Canada, Argentina,
China, India, Australia, and South Africa suggest that single transgene
products in standard cultivar backgrounds are not a recipe for
sustainable pest management. Instead, a much more complex approach may
be needed, one which may involve deployment of a combination of
different transgenes in different backgrounds. Under diverse climatic
conditions and cropping systems of tropics, the success in the
utilization of transgenics for pest management may involve decentralized
national breeding programs and several small-scale seed companies. While
several transgenic crops with insecticidal genes have been introduced in
the temperate regions, very little has been done to use this technology
for improving crop productivity in the harsh environments of the
tropics, where the need for increasing food production is most urgent.
There is a need to develop appropriate strategies for deployment of
transgenics for pest management, keeping in view the pest spectrum
involved, and the effects on nontarget organisms in the ecosystem.

Document Number: 415

Cassava (Manihot esculenta, Crantz) roots are the primary source of
calories for more than 500 million people, the majority of whom live in
the developing countries of Africa. Cassava leaves and roots contain
potentially toxic levels of cyanogenic glycosides. Consumption of
residual cyanogens (linamarin or acetone cyanohydrin) in incompletely
processed cassava roots can cause cyanide poisoning. Hydroxynitrile
lyase (HNL), which catalyses the conversion of acetone cyanohydrin to
cyanide, is expressed predominantly in the cell walls and laticifers of
leaves. In contrast, roots have very low levels of HNL expression. We
have over-expressed HNL in transgenic cassava plants under the control
of a double 35S CaMV promoter. We show that HNL activity increased more
than twofold in leaves and 13-fold in roots of transgenic plants
relative to wild-type plants. Elevated HNL levels were correlated with
substantially reduced acetone cyanohydrin levels and increased cyanide
volatilization in processed or homogenized roots. Unlike acyanogenic
cassava, transgenic plants over-expressing HNL in roots retain the
herbivore deterrence of cyanogens while providing a safer food product.

Document Number: 416

Cassava (Manihot esculenta, Crantz) roots are the primary source of
calories for more than 500 million people, the majority of whom live in
the developing countries of Africa. Cassava leaves and roots contain
potentially toxic levels of cyanogenic glycosides. Consumption of
residual cyanogens (linamarin or acetone cyanohydrin) in incompletely
processed cassava roots can cause cyanide poisoning. Hydroxynitrile
lyase (HNL), which catalyses the conversion of acetone cyanohydrin to
cyanide, is expressed predominantly in the cell walls and laticifers of
leaves. In contrast, roots have very low levels of HNL expression. We
have over-expressed HNL in transgenic cassava plants under the control
of a double 35S CaMV promoter. We show that HNL activity increased more
than twofold in leaves and 13-fold in roots of transgenic plants
relative to wild-type plants. Elevated HNL levels were correlated with
substantially reduced acetone cyanohydrin levels and increased cyanide
volatilization in processed or homogenized roots. Unlike acyanogenic
cassava, transgenic plants over-expressing HNL in roots retain the
herbivore deterrence of cyanogens while providing a safer food product.

Document Number: 417

Genetically modified (GM) crops have been in use commercially around the
world for almost a decade. This review covers the successes and failures
of GM crop varieties in that time, the current status of GM crop
adoption and the traits that are being used. It also describes some of
the GM crops that might come on to the market in the next decade. The
barriers in the way of GM crop development in Europe, including consumer
hostility, the difficulty in gaining official approval and
discriminatory labelling laws are discussed.

Document Number: 418

The current status of cassava Begomoviruses, the most serious constraint
to the production of cassava, a major staple food crop in sub-Saharan
Africa, is reviewed in relation to their distribution, effects,
etiology, and epidemiology. It is concluded that control of the diseases
would continue to depend on integrated management involving cultural
practices and use of resistant cultivars. Current trends in diagnosis
and control, including the production of transgenic plants, selection
for resistance in cassava via molecular markers, and the determination
of resistant profiles of cassava genotypes to a range of virus variants,
are also discussed. (C) 2004 by The Haworth Press, Inc. All rights
reserved.

Document Number: 419

Analytical techniques to track plant genes in the environment and the
food chain are essential for environmental risk assessment, government
regulation and production and trade of genetically modified (GM) crops.
Here, I review laboratory techniques to track plant genes during
pre-commercialization research on gene flow and post-commercialization
detection, identification and quantification of GM crops from seed to
supermarket. At present, DNA- and protein-based assays support both
activities but the demand for fast, inexpensive, sensitive methods is
increasing. Part of the demand has been generated by stringent food
labeling and traceability regulations for GM crops. The increase in GM
crops, changes in GM crop design, evolution of government regulations
and adoption of risk-assessment frameworks will continue to drive
development of analytical techniques.

Document Number: 420

We report the identification and biotechnological utility of a plant
gene encoding the tocopherol (vitamin E) biosynthetic enzyme
2-methyl-6-phytylbenzoquinol methyltransferase. This gene was identified
by map-based cloning of the Arabidopsis mutation vitamin E pathway
gene3-1 (vte3-1), which causes increased accumulation of
delta-tocopherol and decreased gamma-tocopherol in the seed. Enzyme
assays of recombinant protein supported the hypothesis that At-VTE3
encodes a 2-methyl-6-phytylbenzoquinol methyltransferase. Seed-specific
expression of At-VTE3 in transgenic soybean reduced seed
delta-tocopherol from 20 to 2%. These results confirm that At-VTE3
protein catalyzes the methylation of 2-methyl-6-phytylbenzoquinol in
planta and show the utility of this gene in altering soybean tocopherol
composition. When At-VTE3 was coexpressed with At-VTE4 (gamma-tocopherol
methyltransferase) in soybean, the seed accumulated to >95%
alpha-tocopherol, a dramatic change from the normal 10%, resulting in a
greater than eightfold increase of alpha-tocopherol and an up to
fivefold increase in seed vitamin E activity. These findings demonstrate
the utility of a gene identified in Arabidopsis to alter the tocopherol
composition of commercial seed oils, a result with both nutritional and
food quality implications.

Document Number: 421

The Wavelength Dispersive X-ray fluorescence (WD XRF) determination of
phosphorus in GMO and non GMO food samples is proposed. The tested
materials included commercially available transgenic, unmodified
soya-foods and popular dairy products. The WD XRF method was compared
with the standard molybdenum blue method. Matrix effects were minimised
by using standard reference material. Obtained results were discussed in
respect of the accuracy, statistical parameters and values recommended
by the Recommended Dietary Allowance (RDA). The results indicate the
validity of the proposed methods to routine food analysis. (C) 2003
Elsevier Ltd. 

Document Number: 422

Abiotic stresses, such as drought, salinity, extreme temperatures,
chemical toxicity and oxidative stress are serious threats to
agriculture and the natural status of the environment. Increased
salinization of arable land is expected to have devastating global
effects, resulting in 30% land loss within the next 25 years, and up to
50% by the year 2050. Therefore, breeding for drought and salinity
stress tolerance in crop plants (for food supply) and in forest trees (a
central component of the global ecosystem) should be given high research
priority in plant biotechnology programs. Molecular control mechanisms
for abiotic stress tolerance are based on the activation and regulation
of specific stress-related genes. These genes are involved in the whole
sequence of stress responses, such as signaling, transcriptional
control, protection of membranes and proteins, and free-radical and
toxic-compound scavenging. Recently, research into the molecular
mechanisms of stress responses has started to bear fruit and, in
parallel, genetic modification of stress tolerance has also shown
promising results that may ultimately apply to agriculturally and
ecologically important plants. The present review summarizes the recent
advances in elucidating stress-response mechanisms and their
biotechnological applications. Emphasis is placed on transgenic plants
that have been engineered based on different stress-response mechanisms.
The review examines the following aspects: regulatory controls,
metabolite engineering, ion transport, antioxidants and detoxification,
late embryogenesis abundant (LEA) and heat-shock proteins.

Document Number: 423

Temperature and pH are important parameters in food processing. Their
effect on degradation of plasmid and plant DNA was investigated.
Application of HPLC permitted us to quantitatively distinguish between
covalently closed circular and nicked plasmid DNA. Nicking of plasmid
DNA in buffer increased strongly at decreased pH (99% within 90 min at
65 degreesC/pH 4.0). A virtually identical degradation rate was observed
in tomato serum. Temperature exerted only minor effects (15% within 90
min at 85 degreesC/pH 8.4). The combined effect of the parameters was
additive. Degradation of plant DNA was monitored by determining the
maximal detectable fragment length using polymerase chain reaction. With
transgenic maize DNA in buffer, fragments up to 957 bp remained
detectable for at least 90 min upon application of 65 degreesC at pH
4.0. Soymilk and tofu were produced from genetically modified soybeans.
After boiling (10 min) of raw soymilk, the detectable fragment length
decreased from 1,714 to 1,339 bp. The study of the final tofu product
did not reveal any further DNA degradation. In commercial soy protein
isolate, a rather highly processed food, DNA fragments of at least 714
bp were detected.

Document Number: 424

Large parts of agricultural soil are contaminated with lead (Pb) and
cadmium (Cd). Although most environments are not heavily contaminated,
the low levels observed nonetheless pose a high risk of heavy metal
accumulation in the food chain. Therefore, approaches to develop plants
with reduced heavy metal uptake are important. Recently, many transgenic
plants with increased heavy metal resistance and uptake of heavy metals
were developed for the purpose of phytoremediation. However, to reduce
heavy metal in the food chain, plants that transfer less heavy metals to
the shoot are required. We tested whether an Escherichia coli gene,
ZntA, which encodes a Pb(II)/Cd(II)/Zn(II) pump, could be useful for
developing plants with reduced heavy metal content. Yeast cells
transformed with this gene had improved resistance to Pb(II) and Cd(II).
In Arabidopsis plants transformed with ZntA, ZntA was localized at the
plasma membrane and improved the resistance of the plants to Pb(II) and
Cd(II). The shoots of the transgenic plants had decreased Pb and Cd
content. Moreover, the transgenic protoplasts showed lower accumulation
of Cd and faster release of preloaded Cd than wild-type protoplasts.
These results show that a bacterial transporter gene, ZntA, can be
functionally expressed in plant cells, and that that it may be useful
for the development of crop plants that are safe from heavy metal
contamination.

Document Number: 425

Multiplex PCR procedures were developed for simultaneously detecting
multiple target sequences in genetically modified (GM) soybean (Roundup
Ready), maize (event 176, Bt11, Mon810, T14/25), and canola (GT73,
HCN92/28, MS8/RF3, Oxy 235). Internal control targets (invertase gene in
corn, lectin and beta-actin genes in soybean, and cruciferin gene in
canola) were included as appropriate to assess the efficiency of all
reactions, thereby eliminating any false negatives. Primer combinations
that allowed the identification of specific lines were used. In one
system of identification, simultaneous amplification profiling (SAP),
rather than target specific detection, was used for the identification
of four GM maize lines. SAP is simple and has the potential to identify
both approved and nonapproved GM lines. The template concentration was
identified as a critical factor affecting efficient multiplex PCRs. In
canola, 75 ng of DNA template was more effective than 50 ng of DNA for
the simultaneous amplification of all targets in a reaction volume of 25
muL. Reliable identification of GM canola was achieved at a DNA
concentration of 3 ng/muL, and at 0.1% for GM soybean, indicating high
levels of sensitivity. Nonspecific amplification was utilized in this
study as a tool for specific and reliable identification of one line of
GM maize. The primer cry1A 4-3' (antisense primer) recognizes two sites
on the DNA template extracted from GM transgenic maize containing event
176 (European corn borer resistant), resulting in the amplification of
products of 152 bp (expected) and 485 bp (unexpected). The latter
fragment was sequenced and confirmed to be Cry1A specific. The systems
described herein represent simple, accurate, and sensitive GMO detection
methods in which only one reaction is necessary to detect multiple GM
target sequences that can be reliably used for the identification of
specific lines of GMOs.

Document Number: 426

Mitochondria of cytoplasmic male sterile crop plants contain novel,
chimeric open reading frames. In addition, a number of crops carry
endogenous double-stranded ribonucleic acid (dsRNA). In this study, the
novel proteins encoded by these genetic components were screened for the
presence of potential binding sites (epitopes) of allergy-associated IgE
antibodies, as was previously done with transgenic proteins from
genetically modified crops. The procedure entails the identification of
stretches of at least six contiguous amino acids that are shared by
novel proteins and known allergenic proteins. These stretches are
further checked for potential linear IgE-binding epitopes. Of the 16
novel protein sequences screened in this study, nine contained stretches
of six or seven amino acids that were also present in allergenic
proteins. Four cases of similarity are of special interest, given the
predicted antigenicity of the identical stretch within the allergenic
and novel protein, the IgE-binding by a peptide containing an identical
stretch reported in literature, or the multiple incidence of identical
stretches of the same allergen within a novel protein. These selected
stretches are present in novel proteins derived from oilseed rape and
radish (ORF138), rice (dsRNA), and fava bean (dsRNA), and warrant
further clinical testing. The frequency of positive outcomes and the
sizes of the identical stretches were comparable to those previously
found for transgenic proteins in genetically modified crops. It is
discussed whether novel proteins from conventional crops should be
subject to an assessment of potential allergenicity, a procedure which
is currently mandatory for transgenic proteins from genetically modified
crops.

Document Number: 427

For rapid and simultaneous detection of transgenic elements in
genetically modified (GM) food crops, we explored DNA array technology.
Forty-four oligonucleotide 23- to 31-mers were selected to use in an
array on the basis of melting temperature and sequence specificity.
Selected oligonucleotides consisted of DNA fragments corresponding to
structural and regulatory elements and selectable markers used in
developing transgenic crops, such as potato. Other oligonucleotides
represented endogenous genes from potato to serve as positive controls
and from heterologous crops, such as soybean and canola, to serve as
negative controls. Amino-terminated oligonucleotides were hand-spotted
on activated nylon membrane with a commercial spotting device. Target
DNA was isolated from foliage of transgenic and nontransgenic crops,
including potato, and labeled with digoxigenin-dUTP by random priming
following restriction digestion to reduce DNA fragment size.
Hybridization signals were visualized by an alkaline phosphatase
anti-DIG-Fab conjugate and the chemiluminescent substrate, CDP-star. We
detected the presence or absence of transgenic elements in transgenic
and nontransgenic potato samples. Preliminary studies demonstrated that
more specific and sensitive hybridization signals were generated from an
oligonucleotide probe array than from a PCR product array. We anticipate
that oligonucleotide probe arrays will be useful for regulatory
monitoring of transgenic events.

Document Number: 428

Insect resistant Bt 176 maize has been developed by genetic modification
to resist European borer infection. In the present investigation, the
experiment was conducted to determine the effect of feeding a new hybrid
of Bt 176 maize (NX 6262-Bt 176) on general health condition and
performance of broiler chickens. Maize grains and diets were subjected
to proximate analysis. Amino and fatty acids investigation were applied
for both maize grains before used. To evaluate the degradation of NX
6262 - Bt 176 maize DNA and its metabolic fate in broiler blood, muscles
and organs. One-day-old male broilers were fed ad libitum on either an
experimental diet containing NX 6262- Bt 176 or a control diet
containing the non-modified maize grains for 35 days. Feed consumption
and body weight were recorded weekly during the experimental period. All
chickens were subjected to nutritional evaluation period at day 20 of
age for 5 successive days, to calculate the percentage of apparent
digestible nutrients in both diets. At day 35 samples were collected at
several intervals after feed withdrawal. Prior to slaughter blood
samples were collected from all birds by heart puncture to prevent DNA
cross contamination. Samples from pectoral and thigh muscles, liver,
spleen, kidney, heart muscle, bursa and thymus glands were collected.
Digesta from different sections of the gastrointestinal tract (GIT) were
collected as well. Packed cell volume (PCV) and some serum parameters
were investigated. There were no significant differences between control
and experimental group concerning chemical composition of feeds,
apparent digestible nutrients, and all performance parameters measured
(P > 0.05). Furthermore, there were no differences in the PCV and the
analysed serum parameters between the control and experimental group.
The results of maize DNA digestibility showed that the new variety takes
the normal physiological passage along broiler GIT similar to the
conventional line. In addition, Bt 176 maize DNA appears to be partially
degraded in different parts of GIT comparable to the DNA of the control
maize line. Results of the metabolic fate of maize DNA in broiler blood,
muscles and organs indicated that only short DNA fragments (199 bp)
derived from the plant chloroplast gene could be detected in the blood,
skeletal muscles, liver, spleen and kidney, which disappeared after
prolongation the fasting time. In heart muscle, bursa of Fabricius and
thymus, no plant chloroplast DNA was found. Bt gene specific constructs
from Bt 176 maize were not detected in any investigated blood or tissue
samples.

Document Number: 429

Over the next two decades, aquaculture is expected to contribute more to
the global supply of fish for food use and thus further help to reduce
global poverty and food insecurity. One major challenge for aquaculture
production is a future stable, predictable and high quality feed supply.
Marine oils represent 40% of today's feed, but an increased price and
reduced availability is expected to cause a demand for alternative oil
resources in the near future. In consequence, the use of vegetable oils
as feed for farmed carnivorous cold-water fish is increasing. Although
beneficial in some respects, this requires improved fatty acid
composition in plants such as soybean or rapeseed to meet the
nutritional demands of farmed fish. Plants might be adapted to meet
these needs by the use of functional genomics. Although most genes
encoding enzymes of storage lipid biosynthesis have been identified and
cloned, fatty acid regulation at the molecular level is not fully
understood. Potential replacement of marine resources with plant
ingredients demands extensive multidisciplinary efforts. Combinations of
basic understanding of gene function, transgene integration and
expression, gene interactions, fatty acid metabolism in plants and
animals and finally public acceptance have to be gained. Transgenic
plants with increased amounts of 18:1 n-9 (oleic acid) and 18:3 n-3
(alpha-linolenic acid) fatty acids in the seed olesomes. and extensions
of these into longer fatty acids (e.g. 20:5 n-3 and 22:6 n-3) using seed
specific promoters would improve rapeseed (Canola) quality for use as
fish feed. (C) 2003 Elsevier Science Ireland Ltd. 

Document Number: 430

Successful incorporation of polyunsaturated fatty acids (PUFA) into
foods requires stabilised PUFA protected from oxidation and from
undesirable interactions with other ingredients. PUFA may be introduced
into foods by enrichment with PUFA rich ingredients such as fish oil or
algal oil, by manipulation of feed for animals for production of omega-3
rich milks, eggs or meat products, and by alteration of biosynthetic
pathways of oil-bearing plants to modify the fatty acid compositions.

Document Number: 431

Cassava (Manihot esculenta Crantz.) is the major source of calories for
subsistence farmers in sub-Saharan Africa. Cassava, however, contains
potentially toxic levels of the cyanogenic glucoside, linamarin. The
cyanogen content of cassava foods can be reduced to safe levels by
maceration, soaking, rinsing and baking; however, short-cut processing
techniques can yield toxic food products. Our objective was to eliminate
cyanogens from cassava so as to eliminate the need for food processing.
To achieve this goal we generated transgenic acyanogenic cassava plants
in which the expression of the cytochrome P450 genes (CYP79D1 and
CYP79D2), that catalyze the first-dedicated step in linamarin synthesis,
was inhibited. Using a leaf-specific promoter to drive the antisense
expression of the CYP79D1/CYP79D2 genes we observed up to a 94%
reduction in leaf linamarin content associated with an inhibition of
CYP79D1 and CYP79D2 expression. Importantly, the linamarin content of
roots also was reduced by 99% in transgenic plants having between 60 and
94% reduction in leaf linamarin content. Analysis of CYP79D1/CYP79D2
transcript levels in transgenic roots indicated they were unchanged
relative to wild-type plants. These results suggest that linamarin is
transported from leaves to roots and that a threshold level of leaf
linamarin production is required for transport.

Document Number: 432

Phytopharming, the production of protein biologicals in recombinant
plant systems, has shown great promise in studies performed over the
past 13 years. A secretory antibody purified from transgenic tobacco was
tested successfully in humans, and prevented bacterial re-colonization
after topical application in the mouth. Rapid production of
patient-tailored anti-lymphoma antibodies in recombinant
Tobamovirus-infected tobacco may provide effective cancer therapy, Many
different candidate vaccines from bacterial and viral sources have been
expressed in transgenic plants, and three human clinical trials with
oral delivery of transgenic plant tissues have shown exciting results.
The use of crop plants with agricultural practice could allow cheap
production of valuable proteins, while providing enhanced safety by
avoidance of animal viruses or other contaminants. However development
of this technology must carefully consider the means to ensure the
separation of food and medicinal products when crop plants are used for
phytopharming.

Document Number: 433

Plants are a major source of vitamins in the human diet. Due to their
significance for human health and development, research has been
initiated to understand the biosynthesis of vitamins in plants. The
pathways that are furthest advanced in elucidation are those of
provitamin A, vitamin C and vitamin E. There is little knowledge about
the regulation, storage, sink and degradation of any vitamin made in
plants, or the interaction of vitamin biosynthetic pathways with other
metabolic pathways. Researchers as well as life science companies have
endeavoured to manipulate levels of vitamins in order to create
functional food with enhanced health benefits, and even with the goal of
achieving levels worth extracting from plant tissues. Thus far,
metabolic engineering has resulted in transgenic plants that contain
elevated levels of provitamin A, vitamin C and E, respectively.
Additional research is necessary to identify all relevant target genes
in order to further improve and tailor plants with elevated vitamin
contents at will.

Document Number: 434

Mercury is one of the most hazardous heavy metals and is a particular
problem in aquatic ecosystems, where organic mercury is biomagnified in
the food chain. Previous studies demonstrated that transgenic model
plants expressing a modified mercuric ion reductase gene from bacteria
could detoxify mercury by converting the more toxic and reductive ionic
form [Hg(II)] to less toxic elemental mercury [Hg(0)]. To further
investigate if a genetic engineering approach for mercury
phytoremediation can be effective in trees with a greater potential in
riparian ecosystems, we generated transgenic Eastern cottonwood (Populus
deltoides) trees expressing modified merA9 and merA18 genes. Leaf
sections from transgenic plantlets produced adventitious shoots in the
presence of 50 mum Hg(II) supplied as HgCl,, which inhibited shoot
induction from leaf explants of wild-type plantlets. Transgenic shoots
cultured in a medium containing 25 mum Hg(II) showed normal growth and
rooted, while wild-type shoots were killed. When the transgenic
cottonwood plantlets were exposed to Hg(II), they evolved 2-4-fold the
amount of Hg(O) relative to wild-type plantlets. Transgenic merA9 and
merA18 plants accumulated significantly higher biomass than control
plants on a Georgia Piedmont soil contaminated with 40 p.p.m. Hg(II).
Our results indicate that Eastern cottonwood plants expressing the
bacterial mercuric ion reductase gene have potential as candidates for
in situ remediation of mercury-contaminated soils or wastewater.

Document Number: 435

Oilseed rape ( Brassica napus) is an important source of oils for
industrial and food processes. Agriculturally it is a useful break crop
in cereal rotations but suffers from competition from broad-leaved weeds
that can be difficult to control. To extend the range and efficacy of
weed control options, herbicide-tolerant oilseed rape cultivars have
been developed through both conventional and transgenic genetic
modification processes. Here we compare the success of these approaches
and discuss the implications of the new herbicide tolerance traits in
agricultural production of oilseed rape. We conclude that the impact of
HT traits on the agri-environment will be dependent on the nature of the
trait and the management strategies under which it is employed. We find
no evidence that the technology used in the production of HT cultivars
has any influence on their performance as crops. In fact transgenic HT
cultivars are simpler to monitor for agricultural and environmental
impact than are conventional HT cultivars.

Document Number: 436

Desiccation has many detrimental effects on the structure and function
of biological membranes and proteins and this molecular damage decreases
the freshness appearance of dehydrated foods. Phospholipid membranes are
destabilised upon water stress by insertion of cellular amphiphiles,
phase transition into the gel phase and membrane fusion. Proteins are
denatured and electron transport chains are perturbed leading to
increased formation of reactive oxygen species which cause irreversible
damage of cellular structures. Cells respond to water stress by
generating defense proteins and metabolites and eventually develop
outstanding desiccation tolerance such as in the case of plant seeds and
pollen, fungal spores, crustacean cysts, etc. The molecular bases for
this remarkable phenomenon are not completely understood but several
important principles have been identified. Three biological systems seem
to act in concert to achieve desiccation tolerance: enzymes involved in
osmolyte synthesis; proteins specialised in desiccation protection of
membranes and proteins (LEA proteins), and antioxidant enzymes and
molecules. Both osmolytes and LEA proteins contribute to stabilisation
of membrane and protein structures by conferring preferential hydration
at moderate desiccation and replacing water at extreme desiccation.
Osmolytes also contribute to osmotic adjustment and act as hydroxyl
radical scavengers. Genetically modified plants with increased
production of these defenses could be useful to improve the quality of
dried food.

Document Number: 437

This study evaluated the effects of including meal from
glyphosate-tolerant (Roundup-Ready(R)) canola (RRC) in barley-based
diets for lambs on apparent digestibility of the diets, growth
performance of the lambs, and carcass quality and composition. Four
isonitrogenous diets were prepared that included canola meal [6.5%, dry
matter (DM) basis] from four different sources (two commercially
available canola blends, COM1 and COM2; a transgenic line, RRC and the
parental non-transgenic line from which RRC was derived, PAR). Apparent
digestibilities of the four diets were determined using eight mature
wethers (67.8 +/- 2.3 kg) in a replicated Latin square with four 21-d
periods. No aspect of digestibility (DM, fibre, or nitrogen balance) was
influenced by canola source. The growth trial involved 60 early-weaned
Arcott lambs (30 ewes; 30 wethers; initial age approximately 2 mo;
initial weight 21.5 +/- 1.0 kg). The lambs were blocked by weight and
gender for assignment to treatments, and fed the diets until reaching or
exceeding 45 kg body weight. Intake of DM was similar among lambs fed
COM1, COM2 and PAR diets, and between PAR and RRC (COM1, COM2 > RRC, P <
0.05). Diet did not affect. (P > 0.05) average daily gain or feed
efficiency. Carcass yield grade was higher (P < 0.05) for COM1 and COM2
diets than for PAR or RRC, although carcass composition did not differ
(P > 0.05) between PAR and RRC. Canola source did not affect (P > 0.05)
meat tenderness, as determined by shear force, drip loss or
intramuscular fat content. Meat colour of RRC-fed lambs did not differ
from that of all other treatment groups. In this study, including canola
meal prepared from glyphosate-tolerant canola did not alter diet
digestibility, feed efficiency, growth performance, carcass
characteristics or meat quality of lambs.

Document Number: 438

Questions regarding the digestive fate of DNA and protein from
transgenic, feed have been raised in regard to human consumption and
commercial trade of animal products (e.g., meat, milk, and eggs) from
farm animals fed transgenic crops. Using highly sensitive,
well-characterized analytical methods, pork loin samples were analyzed
for the presence. of fragments of transgenic and endogenous. plant DNA
and transgenic protein from, animals fed meal prepared from conventional
or glyphosate-tolerant Roundup Ready (RR) soybeans. Pigs were fed diets
containing 24, 19, and 14%, RR or conventional soybean meal during
grower, early-finisher, and late-finisher phases of growth,
respectively, and longissimus muscle samples were collected (12 per
treatment) after slaughter. Total DNA was extracted from the samples and
analyzed by PCR, followed by Southern blot hybridization for the
presence of a 272-bp fragment of the cp4 epsps coding region (encoding
the synthetic enzyme 5-enolpyruvylshikimate-3-phosphate synthase derived
from Agrobacterium sp. strain CP4) and a 198-bp fragment of the
endogenous soybean gene le1 (encoding soy lectin). Using 1 mug of input
DNA per reaction, none of the extracted samples was positive for cp4
epsps or le1 at the limit of detection (LOD) for these PCR/Southern blot
assays. The LOD for these assays was shown to be approximately one
diploid genome equivalent of RR soybean DNA, even in the presence of 10
mug of pork genomic DNA. A 185-bp fragment of the porcine preprolactin
(prl) gene, used as a positive control, was. amplified from all samples
showing that the DNA preparations were amenable to PCR amplification.
Using a competitive immunoassay with an LOD of approximately 94 ng of
CP4 EPSPS protein/g of pork muscle, neither the CP4 EPSPS protein nor
the immunoreactive peptide fragments were detected in loin muscle
homogenates from pigs fed RR soybean meal. Taken together, these results
show that neither small fragments of transgenic DNA nor immunoreactive
fragments of transgenic protein are detectable in loin muscle samples
from pigs fed a diet containing RR soybean meal.

Document Number: 439

Studies on genetically modified (GM) feedstuffs, for poultry (and other
livestock species) have not added any substance to public concerns in
Europe about their safety for human or bird health. The compositions of
maize lines engineered for insect resistance (Bt-maize) or herbicide
tolerance (glyphosate) and herbicide-tolerant soybean have all proved to
be essentially indistinguishable from their conventional counterparts.
Consequently, and not surprisingly, comparative feeding studies with
broilers and layers in which conventional maize (50 to 78%) or soybeans
(27%) were replaced in feeds by transgenic varieties, also have failed
to show differences of any significance in production parameters. These
data indicate that feeding studies with target livestock species
contribute very little to the safety assessment of crops engineered for
input traits that have little or no detectable effect on chemical
composition. However, comparative growth studies made with broiler
chicks, particularly sensitive to any change in nutrient supply or the
presence of toxic elements in their feed, can be used to screen for any
unintended adverse consequence of the recombinant event not detected by
compositional analysis. This does, however, depend on whether the GM
plant can be matched to a parental line or another suitable control and
its suitability for inclusion in broiler diets. The discovery that DNA
fragments from the digestive tract can be found in the tissues of
animals evoked interest in the fate of ingested transgenes. Plant DNA
derived from feed has been detected in the muscle, liver, spleen and
kidneys of broilers and layers, although not in eggs. However, no
fragments of transgenic DNA or its expressed protein have been found to
date in poultry meat or eggs or in any other animal tissues examined.

Document Number: 440

In this study, we used polymerase chain reaction (PCR) to detect
genetically modified Roundup Ready soybean (RRS) in miso. Several
different DNA extraction methods had been tested. The CTAB method
published by Lipp et al. and a commercial kit, Nucleospin Food, were
used, because they had the most appropriate performance for miso sample.
Four pairs of primers specific for the inserted genes and crop
endogenous genes in RRS were applied in PCR. Using these primers, the
method showed a false-negative result for the miso sample during a later
period. When another more sensitive method, nested-PCR had been used, we
obtained false-negative result for the sample in later fermentative
stage. Since PCR and nested-PCR can not yield to positive results, using
these two methods to detect transgenic components in miso is not
efficient.

Document Number: 441

A construct-specific real-time PCR assay was developed for quantitative
detection of LibertyLink in rapeseed by using the junction region
between the 35S promoter and the pat gene. For validation of precision
and accuracy of quantification five external genomic and cloned DNA
calibration standards were constructed. Seeds of non-transgenic rapeseed
were pooled with appropriate numbers of seeds of transgenic rapeseed.
Genomic DNA from transgenic rapeseed was diluted with corresponding
volumes of water or genomic DNA from non-transgenic rapeseed. Plasmids
containing DNA fragments of the construct-specific and a
brassica-specific reference gene were diluted with appropriate volumes
of water and last, corresponding amounts of plasmids containing DNA
fragments of the construct-specific construct were spiked in genomic DNA
from non-transgenic rapeseed. The approximately estimated upper and
lower limits of the quantification of calibration standards constructed
with mixed seeds, plasmids in genomic background and water-diluted
genomic DNA of 206-279 and 21-28 copy numbers were similar, whereas
water-diluted plasmid and mixed genomic DNA standards never reached the
quantification thresholds of 13 and 26 copy numbers, respectively. The
accuracy of quantification for all calibration standards was tested by
calculating the recovery of quantification for reference samples. The
highest accuracy was obtained by using standards of mixed genomic DNA
and plasmids in genomic back-ground. This indicates a significant
influence of background DNA on the quantification of genetically
modified (GM) contaminations in rapeseed. Since construct-specific
plasmids can be produced on a large scale, stored for a long time
without loss of quality and spiked in different DNA backgrounds, plasmid
standards can be used in GM quantification as 'gold standards'.

Document Number: 442

Deficiencies of iron and zinc are common worldwide. Various strategies
have been used to combat these deficiencies including supplementation,
food fortification and modification of food preparation and processing
methods. A new possible strategy is to use biotechnology to improve
trace element nutrition. Genetic engineering can be used in several
ways; the most obvious is to increase the trace element content of
staple foods such as cereals and legumes. This may be achieved by
introduction of genes that code for trace element-binding proteins,
overexpression of storage proteins already present and/or increased
expression of proteins that are responsible for trace element uptake
into plants. However, even very high levels of expression may not
substantially increase the iron and zinc contents unless many atoms of
trace elements are bound per protein molecule. Another possibility is to
introduce a protein that specifically enhances trace element absorption
even in the presence of naturally occurring inhibitors, thus improving
bioavailability. Genetically modifying plants so that their contents of
inhibitors of trace element absorption such as phytate are substantially
reduced is another approach. Increasing the expression of compounds that
enhance trace element absorption such as ascorbic acid is also a
possibility, although this has received limited attention so far. Iron
absorption may be increased by higher ascorbic or citric acid content
but require overexpression of enzymes that are involved in the synthetic
pathways. Finally, a combination of all of these approaches perhaps
complemented with conventional breeding techniques may prove successful.

Document Number: 443

There is a growing interest in producing food plants with increased
amounts of flavonoids because of their potential health benefits.
Tomatoes contain small amounts of flavonoids, most of which are located
in the peel of the fruit. It has been shown that flavonoid accumulation
in tomato flesh, and hence an overall increase in flavonoid levels in
tomato fruit, can be achieved by means of simultaneous overexpression of
the maize transcription factors LC and C1. Fruit from progeny of two
modified lines (2027 and 2059) was selected for a detailed analysis and
individual identification of flavonoids, at different stages of
maturity. Nine major flavonoids were detected in the flesh of transgenic
ripe tomatoes. LC/NMR, LC/MS, and LC/MS/MS enabled us to identify these
as kaempferol-3,7-di-O-glucoside (1),
kaempferol-3-O-rutinoside-7-O-glucoside (2), two
dihydrokaempferol-O-hexosides (3 and 4), rutin (5),
kaempferol-3-O-rutinoside (6), kaempferol-3-O-glucoside (7),
naringenin-7-O-glucoside (8) and naringenin chalcone (9), which were
quantified by HPLC/DAD. All but 5, 6, and 9 were detected in tomato for
the first time. The total flavonoid glycoside content of ripe transgenic
tomatoes of line 2059 was about 10-fold higher than that of the
controls, and kaempferol glycosides accounted for 60% of this.
Kaempferol glycosides comprised around 5% of the flavonoid glycoside
content of ripe control tomatoes (the rest was rutin and naringenin
chalcone). The rutin concentration in both transgenic and control fruits
was similar.

Document Number: 444

Plant genetic engineering is a powerful tool for producing crops
resistant to pests, diseases and abiotic stress or crops with improved
nutritional value or better quality products. Currently over 70
genetically modified (GM) crops have been approved for use in different
countries. These cover a wide range of plant species with significant
number of different modified traits. However, beside the technology used
for their improvement, the common component of most GM crops is the
neomycin phosphotransferase II gene (nptII), which confers resistance to
the antibiotics kanamycin and neomycin. The nptlI gene is present in GM
crops as a marker gene to select transformed plant cells during the
first steps of the transformation process. The use of
antibiotic-resistance genes is subject to controversy and intense
debate, because of the likelihood that clinical therapy could be
compromised due to inactivation of the oral dose of the antibiotic from
consumption of food derived from the transgenic plant, and because of
the risk of gene transfer from plants to gut and soil microorganisms or
to consumer's cells. The present article discusses these possibilities
in the light of current scientific knowledge.

Document Number: 445

To study the regulation of leptin secretion in, sheep, we. infused
glucose (0.32 g/h/kg for 12 h) into GH-transgenic animals (n = 8) that
have chronically high plasma concentrations of,ovine Wand insulin,
bin,low,body condition and low plasma leptin concentrations, and
compared the responses with those in controls (n = 8). In both groups,
the infusion increased plasma 1 concentrations of glucose and insulin
within 1 h and, maintained high levels throughout the infusion. period
(P < 0.0001). Compared with controls, GH-transgenics had higher
concentrations of insulin, IGF-1, GH (all P < 0.0001) and cortisol (P <
0.05), but lower GH pulse frequency (P < 0.0001). Overall, leptin
concentrations, were lower in GH-transgenics than in controls (P <
0.01). A postprandial increase in leptin concentrations was, observed,in
both groups, independently of glucose treatment, after which the values
remained elevated in animals infused. with glucose, but returned to
basal levels in,those infused with saline, independently of transgene
status In both, GH-transgenics and controls, glucose, infusion did not
affect the concentrations of GH, TGF-1, or cortisol. In conclusion,
GH-transgenic and control,sheep show similar responses to glucose
infusion for leptin and other. metabolic hormones, despite differences
between them in body,condition and basal levels of these hormones.
Glucose, inssulin, GH, IGF-1 and cortisol are probably not major factors
in the acute control of leptin secretion. in sheep, although sustained
high concentrations of GH and IGF-1 might reduce adipose tissue mass or
inhibit leptin gene expression. (C) 2002 Elsevier Sciences Inc. All
rights reserved.

Document Number: 446

We report the characterisation of the 3'-integration junction between
host plant DNA and integrated gene-construct of the genetically modified
Bt11 maize, and the successive development and first validation of a
transformation event-specific quantitative TaqMan 5'-nuclease PCR assay
for detection and quantitation of Bt11 maize on the LightCycler. The
transgenic DNA in Bt11 is inserted in a tandem repeated DNA sequence
motif of approximately 180 bp, and we discuss some of the problems this
creates for design, development and validation of event-specific PCR
assays. The limit of detection of the PCR assay was estimated to 10
initial template copies. The limit of quantitation in pure maize
materials was estimated to approximately 40 template copies. In food
samples containing high levels of exogenous DNA, the impact of this DNA
was estimated to increase the limit of quantitation to approximately 100
initial template copies. Because of the low number of template copies
required for detection and quantification, in combination with small
size of the amplicon (similar to100 bp), the assay may be applied to
analyses of raw materials as well as processed food and feed.

Document Number: 447

So far two different strategies for engineering high methionine (Met)
grain legumes were followed separately in several laboratories: a) The
transfer of foreign genes encoding Met-rich proteins, and b) the
engineering of Met biosynthesis pathways. In some cases a down
regulation of the formation of endogenous sulfur-containing compounds
was observed due to the expression of Met-rich foreign proteins. Since
this might result from competition of the foreign protein with
endogenous compounds for limited Met supply both strategies were
combined in the present work. Double transformants of narbon bean (Vicia
narbonensis L.) were generated which express seed-specifically the
Met-rich Brazil nut 2S albumin (BNA) as well as a feed-back insensitive
bacterial aspartate kinase (AK) known to stimulate Met biosynthesis in
transgenic tobacco seeds. In order to produce double transformants a
homozygous transgenic BNA line of narbon bean was either retransformed
with the AK gene or crossed with an AK line. For the first time the
influence of a deregulated AK on amino acids of the aspartate pathway
was studied in seeds of a transgenic legume. Effects of expressing the
foreign genes on inorganic sulphate, free and protein-bound Met and
other amino acids of the aspartate pathway as well as on free
sulphhydryl compounds of mature seeds were analysed. AK lines had 10 to
12 percent and the BNA line 80 percent increased Met in mature seeds.
Double transformants showed additive but not synergistic effects of the
expression of AK and BNA gene on seed Met. In their mature seeds
protein-bound Met reached levels 2.0 to 2.4 times higher than in the
wildtype. The Met level of best line corresponds approximately to the
FAO standard for Met in a nutritionally balanced protein for human food
or for feeding monogastric animals.

Document Number: 448

Results of the CP4EPSPS gene sequence in the transgenic Roundup Ready
soybean, and cloning and expression of CP4EPSPS protein using bacterial
expression system are described. Our sequencing results show that DNA
rearrangement occurred between 3' NOS terminator and the genomic plant
DNA. Expressed CP4EPSPS protein was isolated using simple two steps of
DEAE column, and can be used as a source material for the allergic risk
assessment of newly introduced foreign protein in soybean and for the
production of mono- and/or poly-clonal antibodies.

Document Number: 449

Questions regarding the digestive fate of DNA and protein from
transgenic grain have been raised in regard to human consumption and
trade of animal products (e.g., meat, milk, and eggs) from farm animals
fed transgenic crops. Using highly sensitive, fully characterized
analytical methods, fragments of transgenic and endogenous plant DNA, as
well as transgenic protein, were not detected in chicken breast muscle
samples from animals fed YieldGard Corn Borer Corn event MON 810 (YG).
Total DNA was extracted from breast muscle samples from chickens fed for
42 d with a diet including either 55 to 60% YG grain of 55 to 60%
conventional corn grain. DNA preparations were analyzed by PCR followed
by Southern blot hybridization for the presence of a 211-bp fragment of
the Bacillus thuringiensis (Bt) cry1Ab gene and a 213-bp fragment of the
endogenous corn gene sh2 (encoding ADP glucose pyrophosphorylase). By
using 1 mug of input DNA per reaction, none of the extracted samples was
positive for cry1Ab or sh2 at the limit of detection for these PCR
assays. A 396-bp fragment of the chicken ovalbumin (ov) gene, used as a
positive control, was amplified from all samples showing that the DNA
preparations were amenable to PCR amplification. By using a competitive
immunoassay with a limit of detection of approximately 60 ng of Cry1Ab
protein ' per gram of chicken muscle, neither the Cry1Ab protein nor
immunoreactive peptide fragments were detectable in the breast muscle
homogenates from chickens fed YG grain.

Document Number: 450

The carotenoid, tocopherol and chlorophyll metabolic profiles and
content of a selection of fruits and vegetables found commonly in the
diet, have been determined using a rapid RP-HPLC technique with on-line
PDA detection. Information gathered from the screening of secondary
plant metabolites is vital for the accurate determination of the dietary
intake of these micro-nutrients, and in the development of comprehensive
food tables. Determination of basal levels is also necessary for the
rational engineering of health-promoting phytochemicals in food crops.
In addition this approach can also be applied to the routine screening
of products to determine metabolic differences between varieties and
cultivars, as well as between genetically modified and the corresponding
non-genetically modified tissue. (C) 2003 . All
rights reserved.

Document Number: 451

Plant biotechnology - especially in vitro regeneration and cell biology,
DNA manipulation and biochemical engineering - is already changing the
agricultural scene in three major areas: control of plant growth,
protecting plants against biotic stress, and production of specialty
foods, biochemicals and pharmaceuticals. Plant biotechnology faces
several major challenges in the coming decades: alleviating the hazards
of abiotic stress (especially salinity, drought, and extreme
temperatures), improving pest control, maintenance and improvement of
the environment, improvement of food quality and design of 'specialty
food' using biochemical engineering, and production of biomaterials. Two
parallel research approaches will most likely exist simultaneously in
the near future: the transgenic approach (expression of unique genes and
specific promoters and transcription factors), and the non-transgenic
approach (genomics-assisted gene discovery, marker-assisted selection,
efficient mutations, and clonal agriculture). Drought and salinity are
the most serious threats to agriculture and to the environment in many
parts of the world. Key molecular factors that are being used for
genetic engineering of stress-tolerant plants include: over-expression
of specific transcription factors, characterization of dehydrin
proteins, over-production of osmoprotectants, expression of water
channel proteins and ion transporters, expression and characterization
of molecular chaperones, including a novel boiling-stable
homo-oligomeric SP1 protein. Although molecular breeding is routine in
agriculture, forest-tree species have been left far behind. However, the
increasing demand for wood and its products and the reduction of
available harvestable forests has recently led to the introduction of
several molecular and biotechnological tools into forest-tree research
and improvement. Among these are in vitro propagation, the
identification of molecular markers, and genetic engineering for
specific traits. Achievements today in plant biotechnology have already
surpassed all previous expectations. The full realization and impact of
the new developments depend not only on continued successful and
innovative research and development activities, but also on a favorable
regulatory climate and public acceptance. Plant scientists now have a
central role in society.

Document Number: 452

Plant breeding has been the classical means of manipulating the plant
genome to develop resistant cultivars for controlling plant diseases.
However, genetic engineering provides an entirely new approach.
Although, currently, the area planted with crops genetically modified
for resistance to disease is small compared with that of crops
genetically modified for tolerance to herbicides or resistance to
insects, numerous strategies are being pursued to render plants
resistant to viruses, fungi, bacteria, and nematodes. Risks associated
with the use of transgenic crop plants for agricultural food production
may relate to the use of specific transgenes, marker genes, or gene
control sequences, whereas other concerns address the entire approach of
engineering heterologous genes into plants. Inherent risks can be
mitigated by new and innovative strategies, and transgenic plants can be
produced within a regulatory framework. If genetically modified plants
are to be regulated, it is inevitable that methods will be required to
distinguish between transgenic and nontransgenic plants and their
products. Analytical laboratory tests may be directed toward novel gene
products or the gene construct itself and must take both theoretical and
practical detection limits into consideration. With a judicious
approach, disease-protected transgenic plants may yet prove to be
essential for producing a safe and adequate world food supply without
endangering the environment or jeopardizing plant diversity.

Document Number: 453

The passage and fate of ingested DNA in 48 pigs fed with diets
containing (n = 12) parental or (n = 36) transgenic (Bt) maize were
examined. Pigs were fattened from an initial live weight of 24 kg to
approximately 108 kg. Animals fed transgenic maize were slaughtered in
groups (n = 6) 4, 8, 12, 24, 48 and 72 h after feeding the last
maize-containing diet. Those slaughtered at up to 12 h received no
further feed, while those held for longer prior to slaughter received a
diet in which maize was replaced by barley and wheat. Control animals
were slaughtered at 4 and 8 h. DNA extracted from tissues and gut
contents was examined by PCR for the presence of plant DNA and for any
transgenic material. Recombinant DNA was detectable in the intestinal
contents up to 48 h after the last feeding of a diet containing the
transgenic maize. PCR amplification of plant gene spacers produced
fragments of different sizes, dependent on feed source. The feed source
of rectum samples depended on individual passage rate in the groups and
their restriction analysis showed grain species-specific patterns.
Recombinant or maize-specific DNA was not detectable in tissue samples
of pigs. In contrast, plant DNA fragments were detectable in the
investigated pig tissues.

Document Number: 454

Vitamin-A deficiency (VAD) is a major malnutrition problem in South
Asia, where indica rice is the staple food. Indica-type rice varieties
feed more than 2 billion people. Hence, we introduced a combination of
transgenes using the biolistic system of transformation enabling
biosynthesis of provitamin A in the endosperm of several indica rice
cultivars adapted to diverse ecosystems of different countries. The rice
seed-specific glutelin promoter (Gt-1 P) was used to drive the
expression of phytoene synthase (psy), while lycopene beta-cyclase (Icy)
and phytoene desaturase (crtl), fused to the transit peptide sequence of
the pea-Rubisco small subunit, were driven by the constitutive
cauliflower mosaic virus promoter (CaMV35S P). Transgenic plants were
recovered through selection with either CaMV35S P driven hph (hygromycin
phosphotransferase) gene or cestrum yellow leaf curling virus promoter
(CMP) driven pmi (phophomannose isomerase) gene. Molecular and
biochemical analyses demonstrated stable integration and expression of
the transgenes. The yellow colour of the polished rice grain evidenced
the carotenoid accumulation in the endosperm. The colour intensity
correlated with the estimated carotenoid content by spectro photometric
and HPLC analysis. Carotenoid level in cooked polished seeds was
comparable (with minor loss of xanthophylls) to that in non-cooked seeds
of the same transgenic line. The variable segregation pattern in T-1
selfing generation indicated single to multiple loci insertion of the
transgenes in the genome. This is the first report of using
nonantibiotic pmi driven by a novel promoter in generating transgenic
indica rice for possible future use in human nutrition.

Document Number: 455

Background: Lactoferrin has been Suggested to have many biologic
activities, such as facilitating iron absorption and having
antimicrobial and anti inflammatory effects. In humans, several of these
activities are likely to only be facilitated by human lactoferrin
because they depend on the binding of human lactoferrin to specific
receptors. Rice may be a useful vehicle to introduce recombinant human
lactoferrin to infant foods because it has low allergenicity and is
likely to be safer than using microorganisms or transgenic animals.
Methods: Recombinant human lactoferrin was expressed in the rice cell
culture system, and its biologic activity was assessed by iron-binding
and -releasing properties, antimicrobial activity, and binding and
uptake to Caco-2 cells. The authors also compared the stability of
recombinant and native human lactoferrins against heat, low pH and in
vitro digestion.
Results: Biologic activity of rice-expressed recombinant human
lactoferrin was similar to that of native human lactoferrin.
Heat-treated proteins retained their functional activities except with
severe treatment at 100degreesC for 8 seconds, which disturbed the
iron-binding capacity of recombinant human lactoferrin. Both types of
proteins retained their functional activities between pH 2 and 7.4.
After in vitro digestion, 50% of both proteins were detectable by enzyme
linked immunosorbent assay. The remaining native and recombinant
lactoferrins retained antimicrobial and Caco-2 binding and uptake
activities.
Conclusions: The results indicate recombinant human lactoferrin has
stability similar to native human lactoferrin when exposed to thermal
treatment, pH treatment, and in vitro digestion, suggesting it may be
active when added to infant formula.

Document Number: 456

Methylmercury is an environmental pollutant that biomagnifies in the
aquatic food chain with severe consequences for humans and other
animals. In an effort to remove this toxin in situ, we have been
engineering plants that express the bacterial mercury resistance enzymes
organomercurial lyase MerB and mercuric ion reductase MerA. In vivo
kinetics experiments suggest that the diffusion of hydrophobic organic
mercury to MerB limits the rate of the coupled reaction with MerA
(Bizily et al., 2000). To optimize reaction kinetics for organic mercury
compounds, the merB gene was engineered to target MerB for accumulation
in the endoplasmic reticulum and for secretion to the cell wall. Plants
expressing the targeted MerB proteins and cytoplasmic MerA are highly
resistant to organic mercury and degrade organic mercury at 10 to 70
times higher specific activity than plants with the cytoplasmically
distributed wild-type MerB enzyme. MerB protein in endoplasmic
reticulum-targeted plants appears to accumulate in large vesicular
structures that can be visualized in immunolabeled plant cells. These
results suggest that the toxic effects of organic mercury are focused in
microenvironments of the secretory pathway, that these hydrophobic
compartments provide more favorable reaction conditions for MerB
activity, and that moderate increases in targeted MerB expression will
lead to significant gains in detoxification. In summary, to maximize
phytoremediation efficiency of hydrophobic pollutants in plants, it may
be beneficial to target enzymes to specific subcellular environments.

Document Number: 457

Plant sterols and their hydrogenated forms, stanols, have attracted much
attention because of their benefits to human health in reducing serum
and LDL cholesterol levels, with vegetable oil processing being their
major source in several food products Currently sold. The predominant
forms of plant sterol end products are sitosterol, stigmasterol,
campesterol and brassicasterol (in brassica). In this study,
3-hydroxysteroid oxidase from Streptomyces hygroscopicus was utilized to
engineer oilseeds from rapeseed (Brassica napus) and soybean (Glycine
max), respectively, to modify the relative amounts of specific sterols
to stanols. Each of the major phytosterols had its C-5 double bond
selectively reduced to the corresponding phytostanol without affecting
other functionalities, such as the C-22 double bond of stigmasterol in
soybean seed and of brassicasterol in rapeseed. Additionaly, several
novel phytostanols were obtained that are not produced by chemical
hydrogenation of phytosterols normally present ill plants. (C) 2003
. 

Document Number: 458

Risk governance of GM plants and GM food products is presently subject
to heated scientific and public controversies. Scientists and
representatives of the biotechnology industry have dominated debates
concerning safety issues. The public is suspicious with regard to the
motives of scientists, companies, and political institutions involved.
The dilemmas posed are nested, embracing value questions, scientific
uncertainty, and contextual issues. The obvious lack of data and
insufficient information concerning ecological effects call for
application of the Precautionary Principle (PP). There are, however,
divergent opinions among scientists about the relevance of putative
hazards, definition of potential "adverse effects," and whether actions
should be taken to prevent harm. The reliance on the concept of
substantial equivalence in safety evaluation of GM food is equally
controversial. Consequently, value assumptions embedded in a scientific
framework may be a barrier for employment of the PP. One of our major
conclusions is that precautionary GMP usage requires risk assessment
criteria yet undeveloped, as well as broader and more long-term
conceptions of risk, uncertainty, and ignorance. Conflicts of interest
and public participation are other issues that need to be taken into
consideration. GMP governance regimes that are justifiable from a
precautionary and ethical point of view must transcend traditional
scientific boundaries to include alternative scientific perspectives as
well as public involvement.

Document Number: 459

Proteinase inhibitors (PIs), lectin and patatin purified from potato
tubers were tested in no-choice feeding assays with pollen beetle larvae
(Meligethes spp.). The idea was to search for resistance factors
possible to introduce into oilseed rape (Brassica napus) by genetic
engineering. The larval diet was prepared by soaking oilseed rape
anthers in protein solutions of known concentrations. Potato lectin was
the most potent in that it was the only of these proteins that reduced
both larval survival and growth rate, while cysteine, aspartic and
metallo PIs and patatin only reduced larval growth rate. Serine Pis had
no significant effect on larval performance. Subsequently, the effect of
potato lectin was compared to that of lectins from other food or feed
crops, resulting in the following mortality-based ranking of activity:
Con A from jackbean > wheat germ lectin > potato lectin > peanut lectin.
In choice tests, larvae did not discriminate between Con A- and
control-diets. These results suggest that the effect of Con A on larvae
is toxic, not deterrent. Adult response was stronger to Con A than to
potato lectin in no-choice tests, just as it was in larvae. However,
adult survival rate and weight was not affected by Con A but the lectin
significantly reduced adult feeding as well as oviposition rates. A
resistance factor that suppresses adult feeding on flower buds is
important for reduced impact of the pollen beetle on the Brassica
oilseed crop.

Document Number: 460

In general, breeding programmes directed at the improvement of forage
have concentrated on easily measurable phenotypes such as yield,
digestibility, resistance to lodging, etc. Selection programmes have
improved forage production but historically have addressed relatively
few quality considerations. In addition, selection for quality has been
limited by availability of suitable analytical techniques. With the
current emphasis on quality rather than quantity and the desire by the
public for greater understanding about where their food comes from,
quality considerations should be a greater target in future breeding
programmes. This review briefly covers previous improvements in quality
of grazed and silage forages and considers how new technologies might be
employed to realise targets for future improvements. In particular we
address the concept that interactions between rumen micro-organisms and
ingested plant material in the rumen are not static but are in fact
dynamic. This has implications for post-ingestion biology and feed
utilisation.

Document Number: 461

Cassava roots, a major food in Africa, contain cyanogenic glucosides
that may cause toxic effects. Malawian women farmers considered fields
of seemingly similar cassava plants to be mixes of both 'cool' and
'bitter' cultivars. They regard roots from 'cool' cultivars as
non-toxic. Roots of 'bitter' were considered to require extensive
traditional processing done by women to be safe for consumption. But
curiously, these women farmers preferred 'bitter' cultivars since
toxicity confers protection against theft, which was a serious threat to
the food security of their families. We studied how well these farmers
comprehend the effects of genetic variations in cassava when dealing
with cyanogenesis in this complex system. Using molecular markers we
show that most plants farmers identified as belonging to a particular
named cultivar had a genotype typical of that cultivar. Farmers'
ethno-classification into 'cool' and 'bitter' cultivars corresponded to
a genetic sub-division of the typical genotypes of the most common
cultivars, with four-fold higher cyanogenic glucoside levels in the
bitter cultivars. Examining morphology, farmers distinguished genotypes
better than did the investigators when using a standard botanical key.
Undoubtedly, these women farmers grasp sufficiently the genetic
diversity of cassava with regard to cyanogenesis to simultaneously
benefit from it and avoid its dangers. Consequently, acyanogenic
cassava-the breeding of which is an announced good of some cassava
genetic improvement programmes-is not a priority to these farmers.
Advances in molecular genetics can help improve food supply in Africa by
rapid micropropagation, marker assisted breeding and introduction of
transgenic varieties, but can also help to elucidate tropical
small-scale farmers' needs and skills.

Document Number: 462

Overproduction of cellulolytic enzymes through conventional nuclear
transformation approaches posed a major challenge as they can
potentially degrade the cell wall components and thereby affect
transgenic plant growth and development. In this study, we have tested
the possibility to over produce an alkali-thermostable xylanase gene
from Bacillus sp. Strain NG-27 in tobacco plants through chloroplast
expression. Our results showed that the xylanase expression can reach up
to 6% of the total soluble protein, a value comparable to high level
expression reported for several non-cellulolytic proteins in tobacco
chloroplasts. The chloroplast-expressed xylanase retained its activity
even when the leaves were dried under sun or at 42degreesC, offering
flexibility in the agricultural system in transport and storage. The
recombinant enzyme was purified to homogeneity using single step
chromatography with more than 85% recovery. Most importantly, transgenic
plants were indistinguishable from the control untransformed plants in
their morphology, growth and in seed setting. These results open up new
avenues for large scale production of several other industrially useful
cellulolytic enzymes through chloroplast expression.

Document Number: 463

As a consequence of rapid progress in agricultural biotechnology,
increasing amounts of genetically modified (GM) crops have entered the
food and feed chain in recent years. The aim of our study, which
conducted during 2000-2001, was to monitor the incidence of Roundup
Ready(TM) soyabean (RRS) and the GM maize lines Bt176, Bt11, T25, MON810
and StarLink in Egypt. With the exception of StarLink(TM), which was
approved only for feed use, the other constructs were approved for use
as food and/or feed in the US and other countries. Fifty one soyabean
samples and 61 maize samples were randomly collected from different
localities in Egypt. The detection techniques applied were based on
Polymerase Chain Reaction (PCR) using validated, official detection
methods according to Article 35 of the German Federal Foodstuffs Act.
The results of our survey showed that all soyabean samples imported from
Argentina, and 50% of American soyabean samples contained RRS. Of the 20
maize samples imported from USA, 16 contained Bt176, 17 Bt11, 12 MON810,
19 T25 and 9 StarLinkTm. In addition, of the 7 maize samples imported
from Argentina, 4 contained Bt176 and MON810, 5 T25, 6 Bt11 and 2
StarLink(TM). In contrast, all Egyptian local varieties of soyabean and
maize were non transgenic.

Document Number: 464

Plants not only provide food to humans and animals, but also provide a
large number of nonfood products of industrial and chemical importance.
Moreover, they have the ability to purify the air, soil and water on the
earth. Various trials to genetically improve the potential of plants are
actively in progress. Salt-tolerance would be an especially important
ability to bestow upon plants for agricultural and industrial purposes,
because high salinity conditions are ubiquitous on earth and represent
major barriers to growth. Enhancement of resistance against both
hyper-osmotic stress and Na+ toxity is necessary for successful
molecular breeding of salt tolerant plants. Introduction of genes for
osmolyte bio-synthesis is useful to increase hyperosmotic tolerance of
plant cells. It is introduced in this review that genetically engineered
ectoine synthesis results in increased hyperosmotic tolerance of tobacco
cells. High concentrations of Na+ reduce cellular activity by
interfering with vital Na+-sensitive enzymes and by affecting K+
transport. Understanding the regulation of K+ and Na+ homeostasis is
thus indispensable for enhancement of plant Na+ tolerance. My research
group is investigating the Na+ efflux activity of the yeast Na+-ATPase
(Enal) when installed in the plasma membrane of plant cells, and the
rice K+-Na+ co-transporters (HKT) that contribute to the regulation of
K+ and Na+ uptake in root cells.

Document Number: 465

Trees are used to produce a variety of wood-based products including
timber, pulp and paper. More recently, their use as a source of
renewable energy has also been highlighted, as has their value for
carbon mitigation within the Kyoto Protocol. Relative to food crops, the
domestication of trees has only just begun; the long generation time and
complex nature of juvenile and mature growth forms are contributory
factors. To accelerate domestication, and to understand further some of
the unique processes that occur in woody plants such as dormancy and
secondary wood formation, a 'model' tree is needed. Here it is argued
that Populus is rapidly becoming accepted as the 'model' woody plant and
that such a 'model' tree is necessary to complement the genetic resource
being developed in arabidopsis. The genus Populus (poplars, cottonwoods
and aspens) contains approx. 30 species of woody plant, all found in the
Northern hemisphere and exhibiting some of the fastest growth rates
observed in temperate trees. Populus is fulfilling the 'model' role for
a number of reasons. First, and most important, is the very recent
commitment to sequence the Populus genome, a project initiated in
February 2002. This will be the first woody plant to be sequenced. Other
reasons include the relatively small genome size (450-550 Mbp) of
Populus, the large number of molecular genetic maps and the ease of
genetic transformation. Populus may also be propagated vegetatively,
making mapping populations immortal and facilitating the production of
large amounts of clonal material for experimentation. Hybridization
occurs routinely and, in these respects, Populus has many similarities
to arabidopsis. However, Populus also differs from arabidopsis in many
respects, including being dioecious, which makes selling and back-cross
manipulations impossible. The long time-to-flower is also a limitation,
whilst physiological and biochemical experiments are more readily
conducted in Populus compared with the small-statured arabidopsis.
Recent advances in the development of large expressed sequence tagged
collections, microarray analysis and the free distribution of mapping
pedigrees for quantitative trait loci analysis secure Populus as the
ideal subject for further exploitation by a wide range of scientists
including breeders, physiologists, biochemists and molecular biologists.
In addition, and in contrast to other model plants, the genus Populus
also has genuine commercial value as a tree for timber, plywood, pulp
and paper. (C) 2002 Annals of Botany Company.

Document Number: 466

The needs for recognition of novel conventional or transgenic organisms
include protection of patented or Identity Preserved lines, detecting
transgenics and tracing dispersal. We propose simple 'Biobarcodes(TM)'
using universal PCR primers to recognize the universal 'nonsense'
recognition site of all biobarcodes, followed by a variable nonsense
sequence. The proposed sequences are long enough to allow recognition in
spite of mutations, have stop codons to prevent coding, and will not
self anneal. Sequences of PCR-amplifted biobarcodes can be compared to a
universal database.

Document Number: 467

The UN Cartagena Protocol on Biosafety adopted in Montreal, 29 January,
2000 and opened for signature in Nairobi, 15-26 May, 2000 will exert a
profound effect on international trade in genetically modified organisms
(GMOs) and their products. In this paper, the potential effects of
various articles of the Protocol on international trade in GMOs are
analyzed. Based on the present status of imports of GMOs and domestic
research and development of biotechnology in China, likely trends in
imports of foreign GM food and related products after China accedes to
WTO is explored. Also, China's potential countermeasures to control and
regulate imports of GMOs in line with implementation of the Protocol are
discussed. China, in recent times, has increased its food and
agricultural imports substantially from USA and Canada. China imported
soybean 10.42 mill. tons in 2000 and about 15 mill tons in 2001, of
which majority are from USA where GM soybean accounts for 60%. The
plantation of US Monsanto's transgenic Bt cotton was increased to more
than 1 million ha in China in 2001. Though China has paid great
attention to develop biotechnology, it appears to have little scope to
export GMOs and GM products. So China may consider a range of
administrative measures to implement the Cartagena Protocol and to
regulate its import of GMOs and GM agricultural products. Consequently,
the Regulation on Safety of Agri-GMOs was issued on June, 2001 and
followed three detailed rules issued in Jan. of 2002, with a priority to
limit foreign GMOs importing by safety certification and labeling
system. These were outlined taking into account policies adopted in
Western countries such as green barriers to international trade.

Document Number: 468

Immediate-type reactions to articles made from natural rubber latex are
collectively referred to as latex allergy. Some latex-sensitized
patients also experience allergic reactions to various fruits and
vegetables. This phenomenon is called latex-fruit syndrome. Recent
research revealed that several proteins related to the defense
mechanisms of plants are responsible for latex allergy and the
accompanying cross-reactivity. Protective systems of higher plants are
relatively conserved in the course of evolution, and therefore
homologous defense-related proteins are produced regardless of the
species. Once sensitized by such a conserved protein, the patient is
expected to cross-react to taxonomically unrelated plants. Remarkably,
cross-reactive antigens in the causative foods are generally sensitive
to heat and digestive enzymes. Per-oral sensitization is not commonly
established by such food antigens. They are instead considered to elicit
allergic reactions in latex-sensitized patients based on their
cross-reactivity to the corresponding sensitizers. Defense-related
proteins in higher plants are agriculturally useful, and various
environmental stresses up-regulate their production. These proteins are
also suspected to be relevant to the increasing allergies to plants.

Document Number: 469

The world population is expanding rapidly and will likely be 10 billion
by the year 2050. Limited availability of additional arable land and
water resources, and the declining trend in crop yields globally make
food security a major challenge in the 21st century. According to the
projections, food production on presently used land must be doubled in
the next two decades to meet food demand of the growing world
population. To achieve the required massive increase in food production,
large enhancements in application of fertilizers and improvements of
soil fertility are indispensable approaches. Presently, in many
developing countries, poor soil fertility, low levels of available
mineral nutrients in soil, improper nutrient management, along with the
lack of plant genotypes having high tolerance to nutrient deficiencies
or toxicities are major constraints contributing to food insecurity,
malnutrition (i.e., micronutrient deficiencies) and ecosystem
degradation. Plant nutrition research provides invaluable information
highly useful in elimination of these constraints, and thus, sustaining
food security and well-being of humans without harming the environment.
The fact that at least 60% of cultivated soils have growth-limiting
problems with mineral-nutrient deficiencies and toxicities, and about
50% of the world population suffers from micronutrient deficiencies make
plant nutrition research a major promising area in meeting the global
demand for sufficient food production with enhanced nutritional value in
this millennium. Integration of plant nutrition research with plant
genetics and molecular biology is indispensable in developing plant
genotypes with high genetic ability to adapt to nutrient deficient and
toxic soil conditions and to allocate more micronutrients into edible
plant products such as cereal grains.

Document Number: 470

There is an increasing need for high-throughput analyses of plants and
food samples for the presence of specific DNA sequences, e.g. transgenic
contaminations. We developed and optimized conditions for the automated
isolation of DNA from several maize tissues and various edibles
containing maize using the MagNA Pure LC system (Roche Applied Science).
Our results show that the system provided is capable of isolating DNA
from any tested source. Quantification of an endogenous gene by
LightCycler real-time PCR revealed that the DNA is suitable in quality
and quantity for multiple PCR analyses.

Document Number: 471

Tomatoes, a major food source for humans, accumulate a variety of
secondary metabolites including phenolic compounds, phytoalexins,
protease inhibitors, and glycoalkaloids. These metabolites protect
against adverse effects of hosts of predators including fungi, bacteria,
viruses, and insects. Because glycoalkaloids are reported to be involved
in host-plant resistance, on the one hand, and to have a variety of
pharmacological and nutritional properties in animals and humans, on the
other, a need exists to develop a better understanding of the role of
these compounds both in the plant and in the diet. To contribute to this
effort, this integrated review presents data on the history,
composition, and nutrition of tomatoes, with special focus on the
assessment of the chemistry, analysis, composition, nutrition,
microbiology, and pharmacology of the tomato glycoalkaloids comprising
a-tomatine and dehydrotomatine; their content in different parts of the
tomato plant, in processed tomato products, and in wild and transgenic
tomatoes; their biosynthesis, inheritance, metabolism, and catabolism;
plant-microbe relationships with fungi, bacteria, viruses, insects, and
worms; interactions with ergosterol and cholesterol; disruption of cell
membranes; tomatine-induced tomatinases, pantothenate synthetase,
steroid hydroxylases, and cytokines; and inhibition of
acetylcholinesterase. Also covered are tomato-human pathogen
relationships and tomatine-induced lowering of plasma cholesterol and
triglycerides and enhancement of the immune system. Further research
needs in each of these areas are suggested. The overlapping aspects are
discussed in terms of general concepts for a better understanding of the
impact of tomato glycoalkaloids in the plant in general and in food in
particular. Such an understanding can lead to the creation of improved
tomatoes and to improved practices on the farm and in the consumption of
tomatoes.

Document Number: 472

The results of a survey study on food samples produced from genetically
modified soybean and maize collected from the Egyptian market are
presented. Forty samples of soybean and 40 samples of maize products
have been gathered randomly from markets in Cairo and Giza. The genetic
modification was detected by polymerase chain reaction (PCR) using
official detection methods according to 35 of the German Foodstuffs Act.
Samples were investigated for the presence of material derived from the
following genetically modified organisms (GMOs) all of which are
approved for food use in Europe: Roundup Ready(TM) soybean (RRS) and
maize lines Bt176, Btl 1, T25 and MON810. In addition, samples were
examined in qualitative and quantitative analysis for the presence of
material derived from the transgenic maize line Star-Link(TM) (Aventis)
which was approved for animal feed use exclusively in the US. Twenty %
of 40 investigated soy samples contained Roundup Ready(TM) soybean; 15%
of 40 maize samples tested positive for Bt176 and 12.5% positive for
Bt11 maize. Furthermore, the presence of Star-Link(TM) maize could
clearly be demonstrated in four samples mixed with Bt176 and Bt11. The
percentage of StarLink(TM) was less than 1% in quantiative analysis. The
maize lines T25 and MON810 were not detected.

Document Number: 473

Changes in food production and dietary practices are occurring faster
than our understanding of their potential impact on children's health.
Traditionally, pediatric gastroenterologists have studied food with
respect to its nutritive value and digestibility, its influence on
metabolism, its growth-promoting characteristics, and its relationship
to risk and severity of disease. Biotechnology is now expanding the
science of food to include disease prevention and treatment, as well as
the feeding of children on a global scale. Bioengineered ("genetically
modified", or "transgenic") plants were initially developed to enhance
the food supply by increasing crop yields. Such previously developed
transgenic plants are now prevalent worldwide and appear in many
processed food products. The implementation of the technology of genetic
modulation of food plants has led to considerable fear, controversy, and
confusion as the understanding of the technology is poor in the general
population. This review presents an overview of genetically modified
food crops and their potential unique benefits and risks to children's
health. Political, economical, and ecological issues related to
transgenic crops are not discussed.

Document Number: 474

Many countries are developing or implementing regulatory requirements to
monitor for the presence of genetically modified (GM) materials in
seeds, grain, and derived food products using DNA and protein-based
methods. There is no published report on the detection of different GM
transgenes in canola, and this study is aimed at developing qualitative
PCR methods for the three major GM transgenes commercially available in
canola. Primer sequences were generated from GenBank and previously
published information to develop a polymerase chain reaction (PCR)
detection method for Roundup Ready (glyphosate tolerance, GT73 event),
Liberty Link (glufosinate ammonium tolerance, HCN92 event), and BX
(Bromoxynil tolerance, OXY235 event) canola varieties. On using PCR, two
primer pairs for each of the GT73 and HCN92 and one primer pair for
OXY235 amplified specific amplicons for the three GM transgenes. All
three GM transgenes were detected simultaneously by multiplexing the
five primer pairs in a single PCR reaction. Multiplexing of the five
primer pairs for DNA prepared at 1% (one GM seed in 99 non-GM seeds) and
0.5% (one GM seed in 199 non-GM seeds) levels generated the expected DNA
fragments for GT73, HCN92, and OXY235. This information will lay the
groundwork for the development of a quantitative PCR assay for canola
transgenic events.

Document Number: 475

Plants produce secondary metabolites, among others, to protect
themselves against microbial and herbivore attack or UV irradiation.
Certain metabolite classes also function in beneficial interactions with
other organisms. For example, anthocyanin pigments and terpenoid
essential oils have key roles in attraction of flower pollinators.
Secondary metabolites also have direct uses for man. Flavonoids and
terpenoids for example have health-promoting activities as food
ingredients, and several alkaloids have pharmacological activities.
Controlled transcription of biosynthetic genes is one major mechanism
regulating secondary metabolite production in plant cells. Several
transcription factors involved in the regulation of metabolic pathway
genes have been isolated and studied. There are indications that
transcription factor activity itself is regulated by internal or
external signals leading to controlled responses. The aim of this review
is to discuss the regulation of transcription factors involved in
secondary metabolism in plants at gene and protein levels, using
phenylpropanoid and terpenoid indole alkaloid pathways as two
well-studied examples. (C) 2002 . All rights
reserved.

Document Number: 476

Recombinant DNA technology has significantly augmented the conventional
crop improvement, and has a great promise to assist plant breeders to
meet the increased food demand predicted for the 21st century. Dramatic
progress has been made over the past two decades in manipulating genes
from diverse and exotic sources, and inserting them into microorganisms
and crop plants to confer resistance to insect pests and diseases,
tolerance to herbicides, drought, soil salinity and aluminum toxicity;
improved post-harvest quality; enhanced nutrient uptake and nutritional
quality; increased photosynthetic rate, sugar, and starch production;
increased effectiveness of biocontrol agents; improved understanding of
gene action and metabolic pathways; and production of drugs and vaccines
in crop plants. Despite the diverse and widespread beneficial
applications of biotechnology products, there remains a critical need to
present these benefits to the general public in a real and
understandable way that stimulates an unbiased and responsible public
debate. The development, testing and release of agricultural products
generated through biotechnology-based processes should be continuously
optimized based on the most recent experiences. This will require a
dynamic and streamlined regulatory structure, clearly supportive of the
benefits of biotechnology, but highly sensitive to the well being of
humans and environment. (C) 2002 Published by Elsevier Science Ireland
Ltd.

Document Number: 477

In the present study, we explored the expression of human lysozyme in
maturing rice grains. Particle bombardment-mediated transformation was
utilized to deliver the codon-optimized structural gene for human
lysozyme to the callus of rice cultivar Taipei 309. Lysozyme expression
is controlled by the promoter and signal peptide sequence for rice
storage protein Glutelin 1. A total of 33 fertile plants were
regenerated from independent transformation events and 12 of them with
significant expression levels of lysozyme were advanced to further
generations. The transgenes were characterized by PCR and Southern blot
analysis. Segregation analysis indicated a typical Mendelian 3: 1
inheritance, suggesting a single locus or closely linked loci of gene
insertion. The expression levels of lysozyme reached 0.6% of the brown
rice weight or 45% of soluble proteins. Seven transgenic breeding lines
have been selected and followed over six generations. Lysozyme
expression levels were maintained in all generations. Biochemical,
biophysical and functional comparisons of native and recombinant human
lysozyme revealed identical N-terminal sequence, molecular weight, pI
and specific activity. Similar thermal and pH stability was observed for
lysozyme from two sources. Furthermore, similar bactericidal activity
was displayed towards a laboratory strain of E. coli. The possibility of
improving medical and nutritional quality of infant formulas and baby
foods with rice flour or rice extract containing recombinant human
lysozyme is discussed.

Document Number: 478

Monoterpenes, the C-10 isoprenoids, are a large family of natural
products that are best known as constituents of the essential oils and
defensive oleoresins of aromatic plants. In addition to ecological roles
in pollinator attraction, allelopathy and plant defense, monoterpenes
are used extensively in the food, cosmetic and pharmaceutical
industries. The importance of these plant products has prompted the
definition of many monoterpene biosynthetic pathways, the cloning of the
relevant genes and the development of genetic transformation techniques
for agronomically significant monoterpene-producing plants. Metabolic
engineering of monoterpene biosynthesis in the model plant peppermint
has resulted in yield increase and compositional improvement of the
essential oil, and also provided strategies for manipulating flavor and
fragrance production, and plant defense.

Document Number: 479

So far, genetic engineering of plants in the context of insect pest
control has involved insertion of genes that code for toxins, and may be
characterized as the incorporation of biopesticides; into classical
plant breeding. In the context of pesticide usage in pest control,
natural enemies of herbivores have received increasing attention,
because carnivorous arthropods are an important component of insect pest
control. However, in plant breeding programmes, natural enemies of
herbivores have largely been ignored, although there are many examples
that show that plant breeding affects the effectiveness of biological
control. Negative influences of modified plant characteristics on
carnivorous arthropods may induce population growth of new, even more
harmful pest species that had no pest status prior to the pesticide
treatment. Sustainable pest management will only be possible when
negative effects on non-target, beneficial arthropods are minimized. In
this review, we summarize the effects of insect-resistant crops and
insect-resistant transgenic crops, especially Bt crops, from a food web
perspective. As food web components, we distinguish target herbivores,
non-target herbivores, pollinators, parasitoids and predators.
Below-ground organisms such as Collembola, nematodes and earthworms
should also be included in risk assessment studies, but have received
little attention. The toxins produced in Bt plants retain their toxicity
when bound to the soil, so accumulation of these toxins is likely to
occur. Earthworms ingest the bound toxins but are not affected by them.
However, earthworms may function as intermediaries through which the
toxins are passed on to other trophic levels. In studies where effects
of insect-resistant (Bt) plants on natural enemies were considered,
positive, negative and no effects have been found. So far, most studies
have concentrated on natural enemies of target herbivores. However, Bt
toxins are structurally rearranged when they bind to midgut receptors,
so that they are likely to lose their toxicity inside target herbivores.
What happens to the toxins in non-target herbivores, and whether these
herbivores may act as intermediaries through which the toxins may be
passed on to the natural enemies, remains to be studied.

Document Number: 480

There are two common approaches for the detection of Genetically
Modified Organisms (GMOs): DNA based methods, mainly founded on the
Polymerase Chain Reaction (PCR), which detect genetically modified DNA
sequences, and protein based methods, relying on immune assays (e.g.
Enzyme-Linked Immunosorbent Assay, ELISA). The latter detect and measure
levels of proteins expressed by transgenic genes. Official standard
tests, yet to come, will be based on these two methodologies. DNA based
tests are now preferred for their sensitivity and their capability to
detect a wider range of constructs. Protein-based immune assay tests,
although less sensitive, are quicker and require less lab skills.
Collaborative study results put in evidence that PCR tests are generally
well suited for detecting the presence of GMOs on a qualitative basis
(yes/no). Difficulties arise when GMOs have to be quantitatively
identified in food ingredients. Real-Time, or kinetic PCR, points out
quantification and interpretation limits when quantification has to be
done on a very small total DNA amount. An essential requirement for
PCR-based techniques is the knowledge of the GMO-specific DNA sequence
target. Many labs find it difficult to keep up with the rate at which
life science companies are creating new GMOs and the finding of adequate
reference standards to be used as positive analytical controls.

Document Number: 481

Multiplex PCR reactions.,were developed for detecting simultaneously the
CrylA(b) and pat genes from events 176, MON810, BT11, and T25 of
transgenic maize, using only two pairs of primers, one for the CrylA(b)
gene and the other for the pat gene. The Roundup Ready soybean can be
precisely detected by a multiplex PCR reaction using known primers,
amplifying fragments of the NOS and the epsps sequences simultaneously.
Transgenic events such as Roundup Ready soybean and GA21 maize, among
others, can be quantified b real-time PCR using a pair of primers and a
probe specifically designed for annealing to the NOS ending region. As
an alternative to amplifying an endogenous gene, the addition of a
foreign gene in a percentage equal to the required level of detection,
in a parallel reaction, is proposed. The use of hexane to homogenize
large flour samples is suggested.

Document Number: 482

Pest management techniques have evolved over the past 50 years.
Inorganic chemical pesticides were replaced by synthetic organic
chemicals, and now biopesticides constitute a significant part of pest
management technology. Requirements for the regulatory approval of
pesticides changed dramatically in 1996 with the passage of the Food
Quality Protection Act (FQPA). The FQPA directs the U.S. Environmental
Protection Agency (EPA) to make more rigorous and conservative
evaluation of risks and hazards and mandates a special emphasis on the
safety of infants and children. The EPA provides incentives for the
industry to register materials that are designated "reduced risk". The
future for the registrant industry will include continued reduction in
numbers of registrants through mergers and acquisitions. Conventional
chemicals will remain as important pest management components, and the
processes of combinatorial chemistry and high-throughput bioassays will
allow the rapid synthesis and testing of large numbers of candidate
compounds. Biopesticides will become more important tools in pest
management, with microbial pesticides and transgenic crops being likely
to play important crop protection roles. There will be a continuing need
for research-based approaches to pest control.

Document Number: 483

Research on groundnut biotechnology is mainly carried out in the United
States and also through international collaborative programs involving
Icrisat and Cirad. However, despite its high phenotypic variation,
cultivated groundnut shows little molecular polymorphism, The only known
application to DNA marker-assisted selection has been the use of genes
from wild species in crosses with cultivated species. Until quite recent
years, the main results published concerned the development of efficient
plant regeneration and gene transfer techniques. Groundnut is an
important food and cash crop in the semi-arid tropics but drought and
subsequent seeds pre-harvest aflatoxin contamination are the main
constraints to crop productivity and quality. Since genetic engineering
techniques are available and genetic mapping proves unfruitful, research
is now focused on functional genomics. The objective is to provide new
molecular tools to assist breeding programmes for the resistance to
these two complex traits.

Document Number: 484

Some plants can hyperaccumulate metal ions that are toxic to virtually
all other organisms at low dosages. This trait could be used to clean up
metal-contaminated soils. Moreover, the accumulation of heavy metals by
plants determines both the micronutrient content and the toxic metal
content of our food. Complex interactions of transport and chelating
activities control the rates of metal uptake and storage. In recent
years, several key steps have been identified at the molecular level,
enabling us to initiate transgenic approaches to engineer the transition
metal content of plants.

Document Number: 485

The inclusion of genetically modified (GM) foods in the human diet has
caused considerable debate. There is concern that the transfer of
plant-derived transgenes to the resident intestinal microflora could
have safety implications. For these gene transfer events to occur, the
nucleic acid would need to survive passage through the gastrointestinal
tract. The aim of the present study was to evaluate the rate at which
transgenes, contained within GM soya and maize, are degraded in gastric
and small bowel simulations. The data showed that 80 % of the transgene
in naked GM soya DNA was degraded in the gastric simulations, while no
degradation of the transgenes contained within GM soya and maize were
observed in these acidic conditions. In the small intestinal
simulations, transgenes in naked soya DNA were degraded at a similar
rate to the material in the soya protein. After incubation for 30 min,
the transgenes remaining in soya protein and naked DNA were 52 (sem
13.1) % and 34 (sem 17.5) %, respectively, and at the completion of the
experiment (3 h) these values were 5 % and 3 %, respectively. In
contrast to the soya transgene, the maize nucleic acid was hydrolysed in
the small intestinal simulations in a biphasic process in which
approximately 85 % was rapidly degraded, while the rest of the DNA was
cleaved at a rate similar to that in the soya material. Guar gum and
tannic acid, molecules that are known to inhibit digestive enzymes, did
not influence the rate of transgene degradation in soya protein. In
contrast guar gum reduced the rate of transgene degradation in naked
soya DNA in the initial stages, but the polysaccharide did not influence
the amount of nucleic acid remaining at the end of the experiment.
Tannic acid reduced the rate of DNA degradation throughout the small
bowel simulations, with 21 (sem 5.4) % and 2 (sem 1.8) % of the naked
soya DNA remaining in the presence and absence of the phenolic acid,
respectively. These data indicate that some transgenes in GM foods may
survive passage through the small intestine.

Document Number: 486

No nation has ever fabricated or maintained a prosperous food system
based on genetic resources of purely indigenous origin. Remarkably, many
countries now seem ready and almost eager to try such an approach. We
identify four separate components of an emerging regime that are
interacting in ways that should worry everyone concerned with the
development and transfer of plant genetic materials into the South: new
provisions on intellectual property; increased concentration of new
enabling technologies into a few large multinational companies;
heightened anxieties over transgenic crops; and new problems arising
from international agreements. We argue that the solutions now being
discussed in global forums are either infeasible, incomplete, or are
likely to have seriously negative effects. We call instead for creative
new thinking on building human capacity in developing countries, on the
legal status of plant genetic resources, and on public-private
partnerships, especially those in service of the poor. (C) 2002
Published by .

Document Number: 487

Agricultural biotechnology has been widely adopted in agriculture hut is
also the locus of controversy. Questions hake arisen regarding food and
environmental safety. In the LIS. responsibility for ensuring
agricultural and environmental safety is delegated to the USDA and EPA.
respectively. The FDA has primary responsibility for food safety, with
the exception that the EPA has responsibility for the safety of proteins
in plants associated with insect defense mechanisms, The food safety
assessment, whether performed h the FDA or the EPA, requires evaluation
of the safety of 1) the newly added DNA, 2) the safety of the newly
introduced gene product and 3) the overall safety of the balance of the
foods A paradigm called "Substantial Equivalence" guides the assessment.
The principal food safety issues, for new varieties crops are 1)
potential toxicity of the newly introduced protein(s), 2) potential
changes in allergenicity. 3) changes in nutrient composition, 4)
unintended effects giving rise to allergenicity or toxicity and 5) the
safety of antibiotic proteins included with the transgene. All of these
must he taken in the context of the predicted range of dietary
exposures, The evaluation seeks to establish that there is a "reasonable
likelihood of safety" and that new varieties are as safe as or safer
than crop, produced by traditional methods Indeed. after extensive
safety testing and some five bears of experience with such crops in the
marketplace. there is not a single report that would lead an expert food
scientist to question the safety of such transgenic crops now in use.

Document Number: 488

Plant foods can serve as dietary sources of all essential minerals
required by humans, Unfortunately, mineral concentrations are low in
some plants. especially many staple food crop, thus, efforts are
underway to increase the mineral content of these foods as a means to
ensure adequate attainment of dietary, minerals in all individuals,
While these efforts hake included classical breeding. approaches in the
past, it is clear that future progress can be made by utilizing the
tools of biotechnology to effect directed changes in plant mineral
status. Reviewed are the short- and long-distance mineral transport
mechanisms responsible for the root acquisition and whole-plant
partitioning of mineral ions in crop plants. This background is used to
discuss different transgenic strategies with the potential to enhance
mineral content in vegetative and/or reproductive tissues. Due to
various constraints imposed by plant transport systems on whole-plant
mineral movement. it is argued that modifications designed to increase
the supply of minerals to edible organs should hake the highest chance
for Examples of previous efforts to manipulate plant mineral nutrition
through the introduction of no cl transgenes are presented to
demonstrate the utility of these approaches.

Document Number: 489

Objective: Iron deficiency is estimated to affect about 30% of the world
population. iron supplementation in the form of tablets and food
fortification has not been successful in de eloping countries. and iron
deficiency is still the most important deficiency related to
malnutrition, Here we present experiments that aim to increase the iron
content in rice endosperm and to improve its absorption in the human
intestine by means of genetic engineering,
Methods: We first introduced a ferritin gene from Phaseolus vulgaris
into rice grains, increasing their iron content up to twofold. To
increase iron bioavailability, we introduced a thermo-tolerant phytase
from Aspergillus fumigatus into the rice endosperm, In addition, as
cysteine peptides are considered major enhancers of iron absorption, we
over-expressed the endogenous cysteine-rich metallothionein-like
protein,
Results: The content of cysteine residues increased about sevenfold and
the phytase level in the grains about one hundred and thirty fold,
giving a phytase activity sufficient to completely degrade phytic acid
in a simulated digestion experiment.
Conclusions: This rice, with higher iron content. rich in phytase and
cysteine-peptide has a great potential to substantially improve iron
nutrition in those populations, where iron deficiency I, so widely
spread.

Document Number: 490

Plants are the basis of human nutrition and have been selected and
improved to assure this purpose. Nowadays, new technologies such as
genetic engineering and genomics approaches allow further improvement of
plants. We describe here three examples for which these techniques have
been employed. We introduced the first enzyme involved in fructan
synthesis, the sucrose sucrose fructosyltransferase (isolated from
Jerusalem artichoke), into sugar beet. The transgenic sugar beet showed
a dramatic change in the nature of the accumulated sugar, 90% of the
Sucrose being converted into fructan, The use of transgenic sugar beet
for the production and isolation of fructans will result in a more
efficient plant production system of fructans and should promote their
use in human food. ne second example shows how the over-expression of
the key enzyme of flavonoid biosynthesis could increase anti-oxidant
levels in tomato. Introduction of a highly expressed chalcone isomerase
led to a seventyfold increase of the amount of quercetin glucoside,
which is a strong anti-oxidant in tomato. We were also able to modify
the essential amino acid content of potato in order to increase its
nutritional value. The introduction of a feedback insensitive bacterial
gene involved in biosynthesis of aspartate family amino acids led to a
sixfold increase of the lysine content. Because the use of a bacterial
gene could appear to be controversial, we also introduced a mutated form
of the plant key enzyme of lysine biosynthesis (dihydrodipicolinate
synthase) in potato. This modification led to a 15 times increase of the
lysine content of potato. This increase of the essential amino acid
lysine influences the nutritional value of potato, which normally hits
low levels of several essential amino acids. These three examples show
how the metabolism of primary constituents of the plant cell such as
sugar or amino acids, but also of secondary metabolites such as
flavonoids, can be modified by genetic engineering. Producing fructan, a
soluble fiber, increasing the level of flavonoids, an antioxidant, in
tomato or increasing the level of essential amino acids in potato are
all clear examples of plant genetic modifications with possible positive
effects on human nutrition.

Document Number: 491

Vaccination is a great asset for eradication of infectious diseases in
human, and animals. With the prevalence of antibiotic resistant
bacterial strains and an alarming increase in new and re-emerging
pathogens. the need for vaccination continues to be a high priority, for
mammalian diseases In the last several years, a Nobel approach for
developing improved mucosal subunit vaccines has emerged by exploiting
the use of genetically modified plants. It has been demonstrated that
plant-derived antigens are functionally similar to conventional
vaccine,, and can induce neutralizing antibodies in mammalian hosts.
Using genetically engineered plants for the production of immunogenic
peptides also provides a new approach tor the delivery of a plant-based
subunit vaccine, i.e., oral delivery, provided these immunogenic
peptides are expressed in an edible part of the plant. such as grain or
fruit, Thus, food crops can plan a significant new role in promoting
human health by serving as vehicles for both production and delivery of
vaccines.

Document Number: 492

Recent advances in agricultural biotechnology have highlighted the need
for experimental evidence and sound scientific judgment to assess the
benefits and risks to society. Nutrition scientists and other animal
biologists need a balanced understanding of the issues to participate in
this assessment. To date most modifications to crop plants have
benefited producers. Crops have been engineered to decrease pesticide
and herbicide usage, protect against stressors, enhance yields and
extend shelf life. Beyond the environmental benefits of decreased
pesticide and herbicide application, consumers stand to benefit by
development of food crops with increased nutritional value, medicinal
properties, enhanced taste and esthetic appeal. There remains concern
that these benefits come with a cost to the environment or increased
risk to the consumer. Most U.S. consumers are not aware of-the extent
that genetically modified foods have entered the marketplace. Consumer
awareness of biotechnology seems to have increased over the last decade,
yet most consumers remain confused over the science. Concern over the
impact on the safety of the food supply remains low in the United
States, but is substantially elevated in Europe. Before a genetically
engineered crop is introduced into commerce it must pass regulatory
scrutiny by as many as four different federal regulatory bodies to
ensure a safe food supply and minimize the risk to the environment. Key
areas for more research are evaluation of the nutritional benefits of
new crops, further investigation of the environmental impact, and
development of better techniques to identify and track genetically
engineered products.

Document Number: 493

This paper focuses on the determination of the GMO content of maize and
soybean samples Using real-time PCR, comparing simplex and duplex PCR.
The total DNA content of samples was determined by amplifying part of a
maize gene encoding a lipid transfer protein, or part of a soybean
lectin gene. The transgenic DNA was quantified by amplifying part of the
CaMV 35S promoter. The importance of preparation and conservation of
standards as well Lis the relevance of DNA extraction protocol on the
variability of results are discussed. For the determination of low GMO
content, limitation in the number of copies of the target gene to be
amplified is considered. For samples with a theoretical GMO content of
corresponding to the legal threshold for labelling, the value determined
by duplex real-time PCR ranged from 0.85% to 1.20%. Both real-time
simplex and duplex PCRs allowed identification of GMO free samples
without ambiguity. (C) 2002 . 

Document Number: 494

The detection of genetically modified crops in foodstuff relies on
detection of transgenic DNA or protein material in the sample matrix.
Purified DNA or proteins are used as analytical material for polymerase
chain reaction technologies and immunodiagnostics. Successful sample
preparation is critical to the validity of subsequent analysis. For
routine analysis, a good sample preparation technique should be simple,
safe, and inexpensive while reproducibly generating DNA/protein of
sufficient quality and yield. The suitability of isolated DNA or protein
as an analyte for a detection or characterization technique depends on
amount or concentration, purity, and integrity, each of which may be
influenced by sample matrix and the extraction technique, and, in turn,
may impact the validity of analytical techniques. The key sample
preparation steps of homogenization, pretreatment, extraction, and
purification are discussed as well as typical analytical methods.
Consideration is given to application of these steps for particular
sample matrixes to maximize yield, reduce inhibition effects, and
minimize contamination. The choice of the most appropriate and valid
methods for sample preparation from particular foods is discussed with
respect to DNA analysis. Attention is also given to ease of use, cost,
and generic applicability of the procedures.

Document Number: 495

According to European Commission (EC) Regulation 1139/98, foods and food
ingredients that are to be delivered to the final consumer in which
either protein or DNA resulting from genetic modification is present,
shall be subject to additional specific labeling requirements. Since
1994, genetically altered tomatoes, squash, potatoes, canola, cotton,
and soy have been on the market. Recently, insect-resistant and
herbicide-tolerant maize varieties have been introduced. Soy and maize
are 2 of the most important vegetable crops in the world. During the
past 4 years, both protein- and DNA-based methods have been developed
and applied for detection of transgenic soy and maize, and their
derivatives. For protein-based detection, specific monoclonal and
polyclonal antibodies have been developed; for immunochemical detection,
Western blot analysis and enzyme-linked immunosorbent assays are the
most prominent examples. For detection of genetically modified organisms
(GMOs) at the level of DNA, polymerase chain reaction-based methods are
mainly used. For these reactions, highly specific primer sets are
needed. This study compares the principally different methods.
Specificity of methods and the possible risks of false-positive or
false-negative results are considered in relation to sampling, matrix
effects, and food processing procedures. In addition, quantitative
aspects of protein- and DNA-based GM detection methods are presented and
discussed. This is especially relevant as EC regulation 49/2000, which
defines a threshold for an unintentional comingling of 1%, came into
force on April 10, 2000.

Document Number: 496

Consequential to the implementation of European Commission (EC)
Regulation 1139/98, EC Regulation 49/2000, and EC Regulation 50/2000 has
been the need to measure accurately the levels of the genetically
modified (GM) species Roundup Ready Soya and Bt 176 Maize that are
present in food. Analytical methods to detect and quantitate these
transgenic species have received much attention particularly with
respect to the deminimus threshold of 1% for their presence in materials
derived from non-GM identity-preserved (IP) supplies. The relative
advantages and limitations of threshold analysis by double-competitive
polymerase chain reaction (PCR) and quantitative real-time PCR are
discussed in their application to the quantitative analysis of processed
foods. Consideration is also given to other factors involved in the
analyses that affect the performance of quantitative procedures, and to
the many uncertainties involved in the precision of a reported
analytical result.

Document Number: 497

Pectin is a class of complex cell wall polysaccharides with multiple
roles during cell development. Assigning specific functions to
particular polysaccharides is in its infancy, in part, because of the
limited number of mutants and transformants available with modified
pectic polymers in their walls. Pectins are also important polymers with
diverse applications in the food and pharmaceutical industries, which
would benefit from technology for producing pectins with specific
functional properties. In this report, we describe the generation of
potato (Solamon tuberosum L. cv Posmo) tuber transformants producing
pectic rhamnogalacturonan I (RGI) with a low level of arabinosylation.
This was achieved by the expression of a Golgi membrane-anchored
endo-alpha-1,5-arabinanase. Sugar composition analysis of RGI isolated
from transformed and wild-type tubers showed that the arabinose content
was decreased by approximately 709,, in transformed cell walls compared
with wild type, The modification of the RGI was confirmed by
immunolabeling with an antibody recognizing alpha-1,5-arabinan. This is
the first time, to our knowledge, that the biosynthesis of a plant cell
wall polysaccharide has been manipulated through the action of a
glycosyl hydrolase targeted to the Golgi compartment.

Document Number: 498

The use of plants for production and delivery of vaccine proteins has
shown promise in research conducted during the past decade. Genes
encoding bacterial and viral antigens are faithfully expressed,
processed, and assembled in plant cells to form immunogenic proteins.
Studies in animals and humans showed that ingestion of transgenic potato
containing vaccine proteins caused production of antigen-specific
antibodies in serum and mucosal secretions. Future studies must aim to
improve antigen expression and to develop processes to formulate plant
material for antigen stability and convenient oral delivery.
Furthermore, production methods should ensure the containment of
pharmaceutical crops and prevent contamination of the food supply.

Document Number: 499

The US regulatory review process for novel crops is accomplished jointly
by three federal agencies through a science-based, integrated and
multidisciplinary approach, This process assesses safety for inserted
genes and recipient plants, potential for allergenicity, integrity for
genes, functionality for novel proteins, agronomic characteristics and
environmental safety for novel crops, and composition for key nutrients
and toxicants for crops and important products. To date, more than 50
crops produced through biotechnology have been approved for
commercialization and the vast majority of these have been
compositionally comparable to conventional counterparts for nutrient
content and known toxicants. Experimental data that are required for the
process are clearly defined by a decision-tree approach, and for crops
approved to date, feeding studies using livestock have not been required
as part of the regulatory process. Further, the United States Food and
Drug Administration considers the consumption of DNA to be "Generally
Regarded as Safe." Findings from independently conducted studies have
indicated no differences for nutrient composition, digestibilities, and
animal performance when livestock were fed feedstuffs from conventional
and biotech crops. In addition, in independent studies conducted to
determine whether transgenic DNA and proteins can be detected in animal
products, no transgenic plant-source DNA and proteins have been found in
milk, meat, and eggs. (C) 2002 Elsevier Science B.V. All rights
reserved.

Document Number: 500

Genetically modified food and feed crops have entered the Western
market, and genetically modified animals may follow in the near future.
The issues that are commonly addressed in the assessment of the safety
of genetically modified crops are discussed, as well as the analogous
issues that may arise for genetically modified animals. For safety
assessment, the degree of substantial equivalence of the novel food- or
feed-organism to an appropriate counterpart should be established. Based
upon the differences thus found, a further strategy for the safety
assessment is chosen, This may, for example, take the testing of a novel
food or feed for its potential toxicity and allergenicity. Another issue
is that of the possible transfer of genes from the genetically modified
organism to others. This would be a concern, for example, if the
introduced DNA comprised genes coding for resistance against clinically
important antibiotics. Unanticipated long, term health effects of the
netically modified food or feed can be monitored by postmarket
surveillance. National regulations for food and feed derived from
genetically modified organisms may differ, but they are based upon the
same underlying principles. The regulatory status of genetically
modified animals may differ from that of genetically modified crops, as
the foreign gene in some animals will be regarded a veterinary medicine.
National regulatory experience with transgenic food animals has been
gained in a few cases so far. The FAO/WHO has initiated the
harmonisation of the food safety evaluation of genetically modified
organisms. (C) 2002 Elsevier Science B.V. 

Document Number: 501

Fundamentally, original food products reflect the development of local
rural production systems. Through centuries of tradition such products
have been influenced by environmental diversity and specificity, by
genetic biodiversity, by territorial characteristics and by prevailing
management practices. In the case of processed products human inputs and
influences together with technical innovations have also played a role.
The rhythms of seasonal life have forged most of these products. In
contrast, biotechnologies and genetically moditied organisms are new
words in the vocabulary of food. Consumers' awareness and suspicion of
these recent innovations cannot simply be ascribed to distorted
information, emotion and fear of the unknown as some scientists have
proposed, but they derive from the feeling among consumers that they
have not been consulted. Lack of transparency, traceability and
accountability are the terms commonly used to explain consumers'
reluctance and unwillingness to accept even a modest integration of
genetically modified ingredients into human food and animal feedstuffs.
The birth of transgenic organisms presents a new reality for Europe's
consumers and producers. GM foods result from principles of
industrialization and standardization which is totally contrary to the
production methods that characterise traditional products. These
original products depend mainly on local resources, they respect
territoriality and sustain environmental conservation. But also and
above all, these traditional products satisfy food tastes and habits by
deferring to the gustative preferences of consurners for sharpness in
great variety of natural aromas and flavours. The innovation challenge
should aim to sustain natural variability and not simply imposing global
standardisation and uniformity in the food market. (C) 2002 Elsevier
Science B.V. 

Document Number: 502

Storage proteins account for about 50% of the total protein in mature
cereal grains and have important impacts on their nutritional quality
for humans and livestock and on their functional properties in food
processing. Current knowledge of the structures and properties of the
prolamin and globulin storage proteins of cereals and their mechanisms
of synthesis, trafficking and deposition in the developing grain is
briefly reviewed here. The role of the g uteri proteins of wheat in
determining the quality of the grain for breadmaking and how their
amount and composition can be manipulated leading to changes in dough
mixing properties is also discussed.

Document Number: 503

A possible carry over of foreign food DNA into the body after
consumption was examined. After feeding pigs with conventional and
recombinant (Bt-) maize, different body samples were investigated using
DNA-extraction followed by PCR procedures to detect chloroplast genes of
different length (199 bp and 532 bp), a maize-specific gene (zein) and a
specific transgene present in Bt-maize (cryIa). Initially, a
time-dependent degradation of feed DNA in the gastrointestinal tract of
pig's was analysed within the juices from stomach and three parts of the
small intestine (duodenum, jejunum, ileum). Subsequently, a possible
transfer of residual chloroplast specific DNA as well as recombinant
Bt-maize DNA fragments into different pig organs (blood, muscle, liver,
spleen and lymph nodes) was examined. The suitability of the introduced
DNA extraction procedure was verified through amplification of a
universal gene (ubiquitin) demonstrating the successful PCR analysis
within a range of 189-417 bp long DNA. Short chloroplast DNA fragments
(199 bp) could be successfully amplified from the intestinal juices of
pigs up to 12 h after the last feeding. In contrast,
chloroplast-specific DNA was not found in any pig organ investigated so
far. Specific gene fragments from the transgene maize (Bt-maize) were
never detected in any pig sample.
A field study examining supermarket poultry samples (leg, breast and
wing muscle, stomach) led to frequent detections of the short
chloroplast DNA fragment (199 bp). Furthermore, faint signals for the
maize specific zein gene fragment were detected in these poultry
tissues. Additional PCR examinations using unhatched chicken embryos
provided the first indication that neither chloroplast nor maize genes
are present endogenously within the wild-type poultry genome. Therefore,
a transient transfer of short forage DNA into most poultry organs can be
suspected.

Document Number: 504

Recently, transgenic potato plants were created showing underexpression
of the 20R isoform of the 14-3-3 protein. The transgenic plants grown in
tissue culture showed a significant increase in nitrate reductase
activity and a decrease in nitrate level. The transgenic line with the
lowest 14-3-3 quantity was field-trialed (1997-2000) and analyzed. The
reduction in the 14-3-3 protein level consistently resulted in a starch
content increase and in an increase in the ratio of soluble sugars to
starch in the tubers, although the latter was only barely visible. The
determination of amino acid composition in the tubers showed a
significant increase in methionine, proline, and arginine content and a
slight but consistent increase in hydrophobic amino acid and lysine
content in the cells of the transgenic potato plants. We also observed
an increase in the crude protein content, from 19 to 22.1% of the
control value in consecutive years. It is proposed that all of these
changes might have resulted from the downregulation of nitrate reductase
and sucrose phosphate synthase activities by 14-3-3, although other
potential mechanisms cannot be excluded (e.g., an increase in enzyme
protein level). 14-3-3-repressed transgenic plants showed a significant
increase in calcium content in their tubers. It is thus proposed that a
function of the isolated 14-3-3 isoform is in the control of amino acid
synthesis and calcium metabolism. However, the mechanism of this control
is as yet unknown.

Document Number: 505

The manipulation of plant fatty acid metabolism in such ways as to
produce new valuable vegetable oils has recently increased. A number of
factors such as new molecular biology technologies, lipid-deficient
mutant characterizations and the complete sequencing of the A. thaliana
genome have all converged to make engineering fatty acid metabolism
particularly attractive. The main targets involved in the different
genetic manipulations concerned are the genes encoding the enzymes
involved in the triacylglycerol pathways: acyl-ACP thioesterase,
desaturases, transacylases, elongases, and hydroxylases. These genes are
used in the different processes to modify the chain length, the
unsaturation degree and the double-bound position of the carbon chain of
fatty acid. The engineering of oilseeds for industrial uses and human
food has been successfully achieved. One of these successes has been a
transgenic rapeseed able to produce oil containing up to 90% of lauric
acid obtained by introducing genes from foreign plants. The increase of
the saturated fatty acid content of oilseed by genetic manipulation has
also been equally achieved, soybean and rapeseed with high stearate
content have been obtained. The production of unusual fatty acids such
as polyunsaturated, epoxy, hydroxy, acetylenic, petroselinic and erucic
from common crops are also in progress. Finally, metabolism studies
using a high laurate rapeseed fine show that manipulations of the lipid
biosynthesis pathways have limitations and, as a consequence, new
strategies are being elaborated.

Document Number: 506

Genetical modification may immediately change microorganisms, plants and
animals. From these changes, many questions concerning animal nutrition
result as e-g. the nutritional assessment of modified feed (GMP) or feed
additives, the influence of GMP on animal health and the quality of food
of animal origin as well as the fate of recombinant plant DNA and
protein in animals. In 1997, studies (composition, digestibility,
feeding tests) with GMP of the 1(st) generation (without substantial
changes in composition, e.g. Bt corn, Pat corn, Pat sugar beets, Gt
soybeans) were initiated at the Institute of Animal Nutrition of the
Federal Agricultural Research Centre (FAL) Braunschweig, Germany. First
data from these experiments are reported and compared with literature
data. So far, no significant differences in the nutritional assessment
of feed from isogenic and transgenic plants of the 1(st) generation have
been found up to now (e.g. maize grain, maize silage, sugar beet, sugar
beet leaves silage, full fat soybeans). The same applies to the animal
food produced. The so-called substantial equivalence of transgenic
hybrids has been demonstrated. Recombinant plant DNA-constructs were not
detectable in animal tissue samples. Bt corn showed significantly lower
mycotoyin contents than isogenic hybrids. Further studies for
nutritional assessment of GMO of the 2(nd) generation with substantial
changes in composition or nutritional value seem to be necessary.
Finally, proposals for such Studies including unexpected effects and
safety aspects are discussed.

Document Number: 507

Recombinant DNA technology has great potential to enhance and extend the
advantages of conventional plant breeding, and increase the production
and productivity of crops to meet the increasing demand for food and
food products in the future. Judicious application of this technology
provides opportunities for alleviating some of the major constraints to
crop productivity under subsistence fanning conditions in the developing
countries. Considerable progress has been made in developing strategies
for the production and deployment of transgenic crops. However,
biosafety concerns have been raised regarding the deployment and release
of genetically engineered plants. This debate has divided the fanning
and consumer communities over acceptability of genetically modified
foods. There is a need for a thorough investigation regarding the fate
of transgenic plants in the environment, and their interaction with wild
relatives and non-target organisms. The production and release of
transgenic plants should be based on experience and sound scientific
reasoning. The regulatory requirements for deployment of transgenic
crops should be streamlined and harmonized, in order to achieve
sustainable food production, poverty reduction, and environmental
protection in resource-poor countries in the semi-arid tropics.

Document Number: 508

Modern genetic and molecular technologies provide a number of tools that
can be utilized for the development of staple foods with a higher iron
and zinc content and improved bioavailability of these minerals. This
article summarizes current strategies aimed at increasing the
iron-sequestering capacity of the endosperm and improving mineral
bioavailability via in planta synthesis of microbial phytases. A case
study is presented for wheat, and future strategies are discussed
addressing the importance of phytase thermostability.

Document Number: 509

In the early 1990s, China became as the first country to grow a GM crop,
a virus-resistant tobacco variety. In 1994, tomato variety Flavr-Savr
became the first genetically modified food crop to be produced and
Consumed in an industrialized country. The area planted to GM crops shot
Lip from 1.7 million hectares in 1996 to 11 million in 1997 to 27.8
million in 1998 to 39.9 million in 1999 and to more than 44.2 million in
2000. Countries that grow transgenic crops include North and South
America (USA, Canada, Argentina) and China. Sugar beets are being
modified to contain traits such as herbicide-tolerance,
virus-resistance, and fructan biosynthesis. Herbicide-tolerance sugar
beet works similarly to other such crops. The GM sugar beets are most
often tolerance to glufosinate and glyfosate. Sugar beets are
genetically modified to contain the gene encoding for the viral-sheath
protein of BNYVV causes rhizomania among sugar beets. Because sugar
beets are bred to produce high amounts of sucrose, this makes them a
logical choice for fructan production.

Document Number: 510

In this paper, the possibilities of capillary gel electrophoresis (CGE)
to detect transgenic maize in flours are shown. The method is based on
the extraction and amplification by the polymerase chain reaction (PCR)
of a specific DNA fragment from transgenic maize and its subsequent
analysis by CGE with UV detection or laser-induced fluorescence (LIF),
Some useful considerations regarding the optimization of DNA extraction
and amplification conditions are given. Also, a comparison is
established between the two CGE protocols for DNA detection based on
ultraviolet absorption (CGE-UV) and LIF (CGE-LIF). The requirements,
advantages, and limitations of both CGE methods are discussed. To our
knowledge, this is the first paper on the use of CGE-LIF to detect
transgenic food.

Document Number: 511

Biospecific interaction analysis (BIA) was performed using surface
plasmon resonance (SPR) and biosensor technologies to detect genetically
modified Roundup Ready soybean gene sequences. We first immobilized, on
SA sensor chips, single-stranded biotinylated oligonucleotides
containing soybean lectin and Roundup Ready gene sequences, and the
efficiency of hybridization to oligonucleotide probes differing in
length was determined. Second, we immobilized biotinylated PCR products
from nontransgenic soybeans (genomes carrying only the lectin gene), as
well as from genetically modified Roundup Ready soybean, and we injected
the oligonucleotide probes. Furthermore, we used the sensor chips
carrying either lectin and Roundup Ready soybean PCR products or 21-mer
oligonucleotide as probes, and we injected both nonpurified and purified
asymmetric PCR products. The results obtained show that 13 and 15 mer
oligonucleotides are suitable probes to detect genetically modified
Roundup Ready soybean gene sequences (either target oligonucleotides or
PCR products) under standard BIA experimental conditions. By contrast,
when 11 mer DNA probes were employed, no efficient hybridization was
obtained. All the SPR-based formats were found to be useful for
detection of Roundup Ready gene sequences, suggesting that these
procedures are useful for the real-time monitoring of hybridization
between target single-stranded PCR products, obtained by using as
substrates DNA isolated from normal or transgenic soybeans, and
oligonucleotide or PCR-generated probes, therefore enabling a one-step,
nonradioactive protocol to perform detection.

Document Number: 512

This paper is one of several prepared under the project "Food Safety In
Europe: Risk Assessment of Chemicals in Food and Diet" (FOSIE). a
European Commission Concerted Action Programme, organised by the
International Life Sciences Institute. Europe (ILSI). The aim of the
FOSIE project is to review the current state of the science of risk
assessment of chemicals in food and diet, by consideration of the four
stages of risk assessment, that is. hazard identification. hazard
characterisation, exposure assessment and risk characterisation. The
contribution of animal-based methods in toxicology to hazard
identification of chemicals in food and diet is discussed. The
importance of first applying existing technical and chemical knowledge
to the design of safety testing programs for Food chemicals is
emphasised. There is consideration of the presently available and
commonly used toxicity testing approaches and methodologies, including
acute and repeated dose toxicity, reproductive and developmental
toxicity, neurotoxicity, genotoxicity, carcinogenicity, immunotoxicity
and food allergy. They are considered from the perspective of whether
they are appropriate tor assessing food chemicals and whether them are
adequate to detect currently known or anticipated hazards from food.
Gap. in knowledge and future research needs are identified: research or
these could lead to improvements in the methods of hazard identification
for food chemicals. The potential impact of some emerging techniques,
and toxicological issues, on hazard identification for food chemicals,
such as new measurement techniques, the use of transgenic animals,
assessment of hormone balance and the possibilities for conducting
studies in which common human diseases have been modelled. is also
considered. (C) 2002 ILSI. Published by . All rights
reserved.

Document Number: 513

Numerous analytical methods, both qualitative and quantitative, have
been developed to determine reliably the presence and/or the amount of
genetically modified organisms (GMOs) in agricultural commodities, in
raw agricultural materials and in processed and refined ingredients. In
addition to the "classical" methods for DNA and protein analysis, e.g.
polymerase chain reaction and enzyme linked immunosorbent analysis,
certain types of GMO-containing matrices can be profiled by
complementary chemical analysis methods such as chromatography and near
infrared spectroscopy. This review summarises the status of the most
widely used GMO analysis technologies, identifies new areas of
analytical investigation and discusses current needs and future
challenges.

Document Number: 514

Palatinose (isomaltulose, 6-O-alpha-D-glucopyranosyl-D-fructose) is a
structural isomer of sucrose which is produced from sucrose by some
bacterial strains as a reserve material during periods of low carbon
availability. The ability to synthesise palatinose is not only
advantageous for the bacteria but is also of industrial interest since
palatinose is used as a sucrose substitute in food production. To
explore the possibility of palatinose production in plants a recently
isolated sucrose isomerase gene (palI; EC 5.4.99.11) from Erwinia
rhapontici [F. Bornke et al. (2001) J Bacteriol 183: 2425-2430] was
cloned into a plant expression vector between the constitutive 35S CaMV
promoter and the octopine synthase polyadenylation signal. To allow
secretion of the protein into the apoplast the signal peptide of the
potato proteinase inhibitor 11 was N-terminally fused to the palI coding
region. Expression of the protein was verified by northern and western
blot analyses. Efficient secretion of the protein was demonstrated by
palI detection in intercellular fluids. Transgenic plants expressing
palI accumulated high levels of palatinose. As a consequence, transgenic
plants showed severe phenotypic alterations. Young leaves were curled
and developed bleached areas during maturation. Flowers were misshapen
and sterile. Based on nonaqueous fractionation experiments palatinose
was found in several subcellular compartments, indicating limited
membrane transport of the sugar. In contrast to results obtained with
short-term feeding experiments, no evidence for palatinose-mediated
regulation of photosynthetic or defence genes could be obtained in the
transgenic pall-expressing tobacco plants. Based on our results we
conclude that plants can efficiently be used as bioreactors for the
production of palatinose. Furthermore, tissue-specific expression of
pall should allow carbon allocation to specific tissues and/or
cell-types to be modulated.

Document Number: 515

Recent advances in gene isolation, plant transformation, and genetic
engineering are being used extensively to alter metabolic pathways in
plants by tailormade modifications to single or multiple genes. Many of
these modifications are directed toward increasing the nutritional value
of plant-derived foods and feeds. These approaches are based on rapidly
growing basic knowledge, understanding, and predictions of metabolic
fluxes and networks. Some of the predictions appear to be accurate,
while others are not, reflecting the fact that plant metabolism is more
complex than we presently understand. Tailor-made modifications of plant
metabolism has so far been directed into improving the levels of primary
metabolites that are essential for growth and development of humans and
their livestock. Yet, the list of improved metabolites is expected to
grow tremendously after new discoveries in nutritional, medical, and
health sciences. Despite our extensive knowledge of metabolic networks,
many of the genes encoding enzymes, particularly those involved in
secondary metabolism, are still unknown. These genes are being
discovered at an accelerated rate by recent advances in genetic and
genomics approaches. In the present review, we discuss examples in which
the nutritional and health values of plant-derived foods and feeds were
improved by metabolic engineering. These include modifications of the
levels of several essential amino acids, lipids, fatty acids, minerals,
nutraceuticals, antinutritional compounds, and aromas.

Document Number: 516

Foods that contain plant phenolic secondary metabolites that are
antioxidants are getting more attention due to an increase in
experimental data suggesting health-promoting effects when such foods
are ingested as part of a low-fat diet. As the synthesis of phenolic
compounds in plants is known to increase during the defense response to
microorganisms, we investigated specific food-grade microbial
polysaccharides as potential elicitors of mungbean phenolic content.
Mungbean (Vigna radiata) was pretreated with solutions of both xanthan
and gellan gums, as well as commercial yeast extract and purified yeast
glucan (as potential fungal elicitors), and dark-germinated for 1-5
days. Tissue samples were assayed for enzymatic (glucose-6-phosphate
dehydrogenase and guaiacol peroxidase) and antioxidant activity, and for
proline and phenolic content. Microbial polysaccharide treatments were
found to stimulate phenolic content and enzyme activity, as well as
occasional cotyledon pigmentation. In addition, xanthan gum, yeast
extract, and purified yeast glucan treatments stimulated antioxidant
activity. Possible mechanisms linking acid-induced plant growth to
growth induced by food-grade microorganisms (and related
polysaccharides), such as yeasts and acid-producing bacteria, are
hypothesized and discussed.

Document Number: 517

Commercialization of genetically modified organisms (GMOs) have sparked
profound controversies concerning adequate approaches to risk
regulation. Scientific uncertainty and ambiguity, omitted research
areas, and lack of basic knowledge crucial to risk assessments have
become apparent. The objective of this article is to discuss the policy
and practical implementation of the Precautionary Principle. A major
conclusion is that the void in scientific understanding concerning risks
posed by secondary effects and the complexity of cause-effect relations
warrant further research. Initiatives to approach the acceptance or
rejection of a number of risk-associated hypotheses is badly needed.
Further, since scientific advice plays a key role in GMO regulations,
scientists have a responsibility to address and communicate uncertainty
to policy makers and the public. Hence, the acceptance of uncertainty is
not only a scientific issue, but is related to public policy and
involves an ethical dimension.

Document Number: 518

The European Commission's, Quality of Life Research Programme, Key
Action 1 - Health, Food & Nutrition is mission-oriented and aims,
amongst other things, at providing a healthy, safe and high-quality food
supply leading to reinforced consumer confidence in the safety of
European food. Its objectives also include the enhancing of the
competitiveness of the European food supply. Key Action 1 is currently
supporting a number of different types of European collaborative
projects in the area of risk analysis. The objectives of these projects
range from the development and validation of prevention strategies
including the reduction of consumers risks; development and validation
of new modelling approaches; harmonization of risk assessment
principles, methodologies, and terminology; standardization of methods
and systems used for the safety evaluation of transgenic food; providing
of tools for the evaluation of human viral contamination of shellfish
and quality control; new methodologies for assessing the potential of
unintended effects of genetically modified (genetically modified) foods;
development of a risk assessment model for Cryptosporidium parvum
related to the food and water industries; to the development of a
communication platform for genetically modified organism, producers,
retailers, regulatory authorities and consumer groups to improve safety
assessment procedures, risk management strategies and risk
communication; development and validation of new methods for safety
testing of transgenic food; evaluation of the safety and efficacy of
iron supplementation in pregnant women; evaluation of the potential
cancer-preventing activity of pro- and pre-biotic (`synbiotic')
combinations in human volunteers. An overview of these projects is
presented here.

Document Number: 519

Gene technology has the potential to offer many improvements in the
quality and quantity of the world's food supply provided that genuine
concerns regarding safety, environmental impact, information and ethics
are satisfactorily addressed. In this article, some of the benefits as
well as concerns about genetically modified foods are discussed using
examples such as tomatoes, soybeans, corn and rice.

Document Number: 520

The Asian pea pear, Pyrus betulaefolia Bunge, is tolerant to several
disorders in the fruit bodies caused by high humidity and dryness and is
hence widely used as a rootstock for many pear plants suitable for food
sources. We have now successfully transformed P. betulaefolia Bunge by
an Agrobacterium-mediated gene transfer system. Among several wild-type
A. tumefaciens strains examined, only AKE10 induced shoot-forming tumors
at a high frequency on excised cotyledons of P. betulaefolia Bunge
cultured on phytohormone-free medium. Both the nptII (kanamycin
resistance) and GUS (beta -glucuronidase) genes were introduced into the
cotyledons by infection with AKE10 harboring a binary vector, and
regenerated plants were obtained. Southern hybridization and polymerase
chain reaction analyses and histochemical GUS assay indicated that
morphologically normal transformed plants faithfully contained genes
from the vector but not from wild-type oncogenic T-DNA. However,
morphologically abnormal plants additionally possessed the 6b gene
(AK-6b) of AKE10. These results show that non-disarmed A. tumefaciens is
adequate to transfer genes to the Asian pea pear, P. betulaefolia Bunge.

Document Number: 521

The paper reviews briefly the present status and future trends of some
new, emerging food preservation technologies and anticipates the major
lines of developments in relation to functional and transgenic foods.
These topics of innovative food technologies and food products are of
particular importance in view of the European Union's regulation
concerning "novel foods and novel ingredients". The author considers the
obstacles and the needs for more research and other actions such as
safety assessment and balanced communication to the public.

Document Number: 522

International consensus has been reached on the principles regarding
evaluation of the food safety of genetically modified plants. The
concept of substantial equivalence has been developed as part of a
safety evaluation framework, based on the idea that existing foods can
serve as a basis for comparing the properties of genetically modified
foods with the appropriate counterpart. Application of the concept is
not a safety assessment per se, but helps to identify similarities and
differences between the existing food and the new product, which are
then subject to further toxicological investigation. Substantial
equivalence is a starting point in the safety evaluation, rather than an
endpoint of the assessment. Consensus on practical application of the
principle should be further elaborated. Experiences with the safety
testing of newly inserted proteins and of whole genetically modified
foods are reviewed, and limitations of current test methodologies are
discussed. The development and validation of new profiling methods such
as DNA microarray technology, proteomics, and metabolomics for the
identification and characterization of unintended effects, which may
occur as a result of the genetic modification, is recommended. The
assessment of the allergenicity of newly inserted proteins and of marker
genes is discussed. An issue that will gain importance in the near
future is that of postmarketing surveillance of the foods derived from
genetically modified crops. It is concluded, among others that, that
application of the principle of substantial equivalence has proven
adequate, and that no alternative adequate safety assessment strategies
are available.

Document Number: 523

This Botanical Briefing examines the hypothesis that modern agricultural
practice affects natural biotic pollination to the extent that crop
yields suffer. Few staple foods depend on animal pollination and
relatively few other crops are totally dependent on animal pollination.
However, there are many crops of local economic importance whose yield
or quality may be enhanced by good pollinator activity: studies of these
deserve more attention. Amongst those cases already documented,
intensification and habitat loss are the most frequent causes of
pollinator impoverishment reducing crop yield. As yet there is no clear
example of low crop yield resulting from the effect of pesticides or
transgenic plants on pollinators, and only one example involving
herbicides, although each of these agents can affect populations of crop
pollinators. (C) 2001 Annals of Botany Company.

Document Number: 524

A cysteine proteinase inhibitor expressed in potato plants provides the
first demonstration that transgenic resistance to nematodes such as the
potato cyst nematode Globodera pallida can be effective under field
conditions. The highest level of resistance obtained in the field for
one of the four transformed lines of the normally fully susceptible
Solanum tuberosum tuberosum cv. Desiree was 70 +/-9%. The partially
resistant cv. Sante that is currently of commercial use in the UK showed
a resistance of 85 +/-3%. In containment experiments a change in G.
pallida population to one known to be virulent against cv. Sante caused
a significant loss of its resistance from 80 +/-4% to only 51 +/-6%. In
contrast, the resistance of transgenic Desiree was similar for challenge
by the avirulent and virulent populations with values of 68 +/-6% and 71
+/-4%, respectively. Constitutive expression of the cystatin had no
detrimental effect on either number or weight of tubers and the weight
of the haulm for 3 of the 4 lines in the field. The results establish
that transgenic field resistance against G. pallida can be achieved.
Clearly food and environmental safety must be assured before
commercialisation can be contemplated. However, a prima facie case can
be made that the technology is benign and can reduce the use of
environmentally hazardous nematicides.

Document Number: 525

Since duckweed (Lemnaceae family) is a valuable target plant for various
applications including waste water treatment and food purposes, the
expression of homologous or heterologous proteins may offer an extended
range of application. Therefore, the feasability of transformation of
Wolffia columbiana (Lemnaceae) by Agrobacterium tumefaciens-mediated
gene transfer has been elucidated. Several methods were tested to
increase the accessibility of the plant cells for the infecting
Agrobacterium tumefaciens strain LBA4404, harboring p35SGUSINT :
corundum- and gold particle-treatment, vacuum infiltration and
disintegration of the fronds. The resulting overall transformation
efficiency was higher than without any treatment, reaching an average of
3.9% of all fronds showing GUS staining. Induction of Agrobacterium's
vir genes by media conditions as well as the presence of 0.6 M mannitol
during infection resulted in a clear increase of transformation
efficiency. Max. approx. 30%, average 15-20% of fronds showing GUS
staining were obtained both with corundum-treated as well as with vacuum
infiltrated fronds, but transformation pattern was different. Whereas in
the former mainly epidermal and subepidermal cells were transformed, the
latter showed, in addition, transformed inner frond cells, including the
meristematic region. Disintegration of the fronds, followed by vacuum
infiltration, led to whole GUS-stained areas of the frond fragments. The
results as such and the observed transformation patterns will serve as a
basis for offering good conditions either in the in vivo - or the
in-vitro-regeneration of transgenic duckweed fronds.

Document Number: 526

Polymerase chain reaction (PCR) methods are very useful techniques for
the detection and quantification of genetically modified organisms
(GMOs) in food samples. These methods rely on the amplification of
transgenic sequences and quantification of the transgenic DNA by
comparison to an amplified reference gene. Reported here is the
development of specific primers for the rapeseed (Brassica napus)
BnACCg8 gene and PCR cycling conditions suitable for the use of this
sequence as an endogenous reference gene in both qualitative and
quantitative PCR assays. Both methods were assayed with 20 different
rapeseed varieties, and identical amplification products were obtained
with all of them. No amplification products were observed when DNA
samples from other Brassica species, Arabidopsis thaliana, maize, and
soybean were used as templates, which demonstrates that this system is
specific for rapeseed. In real-time quantitative PCR analysis, the
detection limit was as low as 1.25 pg of DNA, which indicates that this
method is suitable for use in processed food samples which contain very
low copies of target DNA.

Document Number: 527

Iron deficiency is estimated to affect about 30% of the world
population, making iron by far the most deficient nutrient worldwide.
Iron supplementation in the form of tablets and food fortification has
not been successful in developing countries, and iron deficiency is
still the most important deficiency related to malnutrition. Here we
present experiments that aim to increase the iron content in rice
endosperm and to improve its absorption in the human intestine by means
of genetic engineering. We first introduced a ferritin gene fi om
Phaseolus vulgaris into rice grains, increasing their iron content up to
twofold. To increase iron bioavailability, we introduced a
thermotolerant phytase from Aspergillus fumigatus into the rice
endosperm. In addition, as cysteine peptides are considered a major
enhancer of iron absorption, we overexpressed the endogenous
cysteine-rich metallothionein-like protein. The content of cysteine
residues increased about sevenfold and the phytase level in the grains
about 130-fold, giving a phytase activity sufficient to completely
degrade phytic acid in a simulated digestion experiment. However, the
fungal protein did not retain its activity after rice cooking. Therefore
a further attempt to specifically reduce the phytic acid in the inner
part of the rice endosperm, the tissue eaten after the milling of rice
grains, is discussed and biochemical analyses of the transgenic rice
seeds are now in progress. (C) 2001 Society of Chemical Industry.

Document Number: 528

In accordance with regulations (EG) No. 1139/98 and (EG) No. 49/2000
food and food ingredients containing genetically modified soy or maize
must be labelled as such from the threshold of 1 %. Presently, there are
no comparable legally regulations for feed with genetically modified
plant components. In this study, 98 different agriculturally used feed
were analysed qualitatively ( 35 LMBG method) and quantitatively
(real-time PCR with LightCycler) with regard to genetically modified DNA
sequences. Altogether, 89 % of samples containing soy also contained
Roundup Ready(TM) soy; in 5.5 % of samples containing maize transgenic
maize was detected. The methods of 35 LMBG applicable for food can also
be applied to feed. The quantitative results obtained with the
LightCycler and commercially available kits clearly showed, that in
contrast to food most of the feed contained transgenic soy material in
excess of 50 %. Therefore, mandatory labelling of genetically modified
ingredients in feed should be legally regulated as soon as possible. The
results of a comparison of different commercially available kits for the
LightCycler demonstrated, that the standardisation and validation of the
quantitative PCR methods of analysis are urgently required.

Document Number: 529

Genetics has played a pivotal role in increasing the world food
production through revolutions in plant and animal sciences. Though the
attention on fisheries has been inadequate but the growing importance of
modem genetic manipulations and biotechnological innovations to
aquaculture has been realized. Recent advances in fish genetics and
molecular biology have provided a suite of useful techniques, which have
several applications in aquaculture. This paper reviews the advancement
in the applications of selection, hybridization, chromosome engineering,
sex control, gene transfer and molecular technologies for enhanced
aquaculture productivity.

Document Number: 530

Current market conditions and food regulations make it necessary for
international and domestic participants in the agrifood industry to
structure supply chains that control the content of genetically modified
(GM) material in their products. Tests to detect and/or quantify GM
components represent an important tool in maintaining such supply
systems. This study assesses the field performance of kits that employ
lateral flow immunotechnology to detect soybeans GM to be resistant to
the herbicide glyphosate. Operators at 23 grain-handling facilities were
paid to conduct analyses on a series of blinded samples containing
defined proportions of conventional and transgenic soybeans. The
observed rate of false positives was 6.7% in an experiment in which the
highest level of GM material was 1% and 22.3% in a second experiment in
which the highest level of GM material was 10%. This difference may be
attributed to increased risk of cross-contamination with the higher
level of transgenic material used in the second study. Samples
containing 0.01% GM material were reported as genetically modified 6.70%
of the time, while samples containing 0.1, 0.5 and 1% GM material were
classified as genetically modified 29.5, 67.7 and 68.2% of the time,
respectively. Thus, the frequencies of false negatives were 93.3, 70.5,
33.3 and 31.8% for samples containing 0.01, 0.1, 0.5 and 1.0% GM
material. Samples containing 10% GM material were correctly reported as
genetically modified in all cases. These results lead to the conclusion
that the kit under study is useful in screening for lots of soybeans
that contain high levels of GM material, but that, as a field tool, it
is not effective in monitoring for GM material at the level of 1.0% or
lower. Statistical and immunochemical analyses were carried out in order
to assess the relative contributions of various factors to the error
observed in these studies. These analyses indicated that limitations in
operator performance, not defects in test kit materials, were the
primary contributors, while sample size may play a secondary role. As
both operator performance and sample size are independent of the
specific characteristics of the test kit used in this study, it appears
justifiable to generalize conclusions obtained here to other similar
test systems.

Document Number: 531

The xylose isomerase genes (xylA) from Thermoanaerobacterium
thermosulfurogenes and Streptomyces rubiginosus were introduced and
expressed in three plant species (potato, tobacco and tomato) and
transgenic, plants were selected on xylose-containing medium. The xylose
isomerase genes were transferred to explants of the target plant by
Agrobacterium-mediated transformation. The xylose isomerase genes were
expressed under the control of the enhanced cauliflower mosaic virus 35S
promoter and the Omega' translation enhancer sequence from tobacco
mosaic virus. In potato and tomato, xylose isomerase selection was more
efficient than the established kanamycin selection. The level of enzyme
activity in the regenerated transgenic plants selected on xylose was
5-25-fold higher than the enzyme activity in control plants selected on
kanamycin. The xylose isomerase system enables transgenic cells to
utilize xylose as a carbohydrate source. In contrast to antibiotic or
herbicide resistance-based systems where transgenic cells survive on a
selective medium but nontransgenic cells are killed, the xylose system
is an example of a positive selection system where transgenic cells
proliferate while non-transgenic cells are starved but still survive.
The results show that a new selection method is established. The xylose
system is devoid of the disadvantages of antibiotic or herbicide
selection, and depends on an enzyme which is already being widely
utilized in specific food processes and that is generally recognized as
safe for use in the starch industry.

Document Number: 532

Internodes, leaves and tuber slices from potato (Solanum tuberosum),
genotype 1024-2, were subjected to particle bombardment. Transient
expression was optimized using the uidA and the luc reporter genes that
encode beta -glucuronidase (GUS) and luciferase, respectively. Stable
transformation was achieved using the neomycin phosphotransferase
(nptII) gene, which confers resistance to the antibiotic kanamycin. The
influence of biological parameters (tissue type, growth period before
bombardment, pre- and post-bombardment osmoticum treatment) and physical
parameters (helium pressure, tissue distance) that are known to possibly
affect stable transformation were investigated. Putative transgenic
plants, which rooted in media containing kanamycin, were obtained from
all of the tissues tested although there were large differences in the
efficiency: internodes (0.77 plants per bombarded explant), microtuber
slices (0.10 plants per bombarded explant) and leaves (0.02 plants per
bombarded explant). Southern blot analysis of putative transgenic plants
confirmed the integration of the transgenes into plant DNA. The results
indicate that an efficient particle bombardment protocol is now
available for both transient and stable transformation of potato
internodal segments, thus contributing to an enhanced flexibility in the
delivery of transgenes to this important food crop.

Document Number: 533

Polyphenol oxidase (PPO) activity of Russet Burbank potato was inhibited
by sense and antisense PPO RNAs expressed from a tomato PPO cDNA under
the control of the 35S promoter from the cauliflower mosaic virus.
Transgenic Russet Burbank potato plants from 37 different lines were
grown in the field. PPO activity and the level of enzymatic browning
were measured in the tubers harvested from the field. Of the tubers from
28 transgenic lines that were sampled, tubers from 5 lines exhibited
reduced browning. The level of PPO activity correlated with the
reduction in enzymatic browning in these lines. These results indicate
that expression of tomato PPO RNA in sense or antisense orientation
inhibits PPO activity and enzymatic browning in the major commercial
potato cultivar. Expression of tomato PPO RNA in sense orientation led
to the greatest decrease in PPO activity and enzymatic browning,
possibly due to cosuppression. These results suggest that expression of
closely related heterologous genes can be used to prevent enzymatic
browning in a wide variety of food crops without the application of
various food additives.

Document Number: 534

Essential oils, synthesized and stored in leaf glandular trichomes, of
the Mentha species are valuated commercially as additives for food
products, cosmetics and pharmaceuticals. Mint production and oil yield
is attenuated by both biotic and abiotic stresses. Consequently, there
is need for development of cultivars with pest resistance and stable oil
quality. Most mint cultivars are natural hybrids vegetatively
propagated. Their sterility impairs the success of conventional breeding
and to date, the application of irradiation mutation techniques have not
resulted in the release of new commercially acceptable cultivars for
widespread use. The paper summarizes the state of mint biotechnology by
discussing advancements related to in vitro culture and genetic
transformation, generation of herbicide resistant plants, and strategies
for enhancing disease resistance and essential oil biosynthesis.

Document Number: 535

Wheat has played an important role in the diets of humans and animals
since earliest recorded history, in large part because of its high
yields and excellent dough-making properties. These characteristics were
obtained through the, at first empirical, and then directed manipulation
of the genetic make-up of this crop species. With the advent of the 20th
century came the appreciation of the role of single- and multiple-gene
inheritance in determining traits of the plant, e.g., yield, grain
quality, and pest resistance. Later in that century came the development
of genetic tools that led to the ability to associate a particular trait
with a specific chromosome and to desirability of access to the gene
pool available in primitive, but compatible, species of wheat. More
recently developed genetic tools have led to RFLP maps, which pinpoint
the precise location of large numbers of genetic loci, and to
expressed-sequence-tag (EST) banks, which provide probes that facilitate
the breeder's efforts at directed genetic manipulation. The genes
identified in the latter approach can also be used in genetic
engineering procedures. Here, one or several genes are stably integrated
into a single plant cell from which, after sustained division, a plant
can be regenerated in which every cell contains the newly introduced
gene(s). Such efforts can make use of a larger gene pool, since the
genes of one organism are often functional in other organisms. Such
approaches have been used in wheat to alter agronomic performance, pest
resistance, and food and feed quality traits, such as dough quality.
Continued progress toward creating improved wheat varieties will depend
on the melding of the technologies of classical and modern genetics.

Document Number: 536

Currently employed transformation systems require selectable marker
genes encoding antibiotic or herbicide resistance, along with the gene
of interest (GOI), to select transformed cells from among a large
population of untransformed cells. The continued presence of these
selectable markers, especially in food crops such as rice (Oryza sativa
L.), is of increasing public concern. Techniques based on DNA
recombination and Agrobacterium-mediated co-transformation with two
binary vectors in a single or two different Agrobacterium strains, or
with super-binary vectors carrying two sets of T-DNA border sequences
(twin T-DNA vectors), have been employed by researchers to produce
selectable marker-free (SMF) transgenic progeny. We have developed a
double right-border (DRB) binary vector carrying two copies of T-DNA
right-border (RB) sequences flanking a selectable marker gene, followed
by a GOI and one copy of the left border sequence. Two types of T-DNA
inserts, one initiated from the first RB containing both the selectable
gene and the GOI, and the other from the second RB containing only the
GOI, were expected to be produced and integrated into the genome. In the
subsequent generation, these inserts could segregate away from each
other, allowing the selection of the progeny with only the GOI. We
tested this vector using two selectable marker genes and successfully
obtained progeny plants in which the second selectable marker gene
segregated away from the first. Using the DRB binary vector system, we
recovered SMF transgenic lines containing a rice ragged stunt virus
(RRSV)-derived synthetic resistance gene in the rice cultivars Jarrah
and Xiu Shui. Approximately 36-64% of the primary transformants of these
cultivars yielded SMF progeny. Among SMF Jarrah transgenic progeny <50%
of plants contained the RRSV transgene. Thus, we have developed an
efficient vector for producing SMF plants that allows straightforward
cloning of any GOIs in comparison with the published 'twin T-DNA
vectors.

Document Number: 537

A comparison was made of eight known and published PCR primer sets to
detect zein, cryIA(b), bla and bar genes and the 35S cauliflower mosaic
virus promoter of transgenic Bt "Maximizer" maize "event 176" (Novartis)
in cooked food products. Transgenic pop corn, polenta and cookies were
used as models. The polenta and cookies were made with different
percentages (100, 1, 0.1% w/w) of transgenic maize flour. The DNA
extracted from samples taken at different times of cooking revealed a
progressive decrease in the average DNA fragment length, which was,
however, still amplifiable by PCR. Of the primer sets tested, Cry 03/04
showed the best performance and led to the detection of transgenic DNA
in final cooked products containing of Bt-maize DNA as low as 0.1%.

Document Number: 538

During the last five years, the global area of transgenic crop
(GM-genetically modified) cultivation increased 25-fold. About 98% of GM
crops are grown in the USA, Argentina and Canada from where they are
sent to many importers of soyabean and maize. The results of feeding
experiments indicate that soyabean meal obtained from herbicide-tolerant
lints and insect-resistant maize are substantially and nutritionally
equivalent to their conventional lines. A higher content of insecticidal
alpha -amylase inhibitors las well as lectins and alkaloids) may
increase plants' resistance to insect attack, as well as the decrease
nutritional value of seeds. Evaluation of the concordance of the
chemical composition of transgenic and conventional crops (i.e.,
verification of substantial equivalence) is not sufficient for proving
the safety of transgenic food. Sub-chronic in vivo experiments as well
as comparison of nutritional equivalence of transgenic and conventional
crops are advisable. Such actions are justified not only by the
possibility of undesirable transgenic effects, but also by the
consumer's right to explicit information on food safety. Without
evaluation of nutritional equivalence, information on GM-food safety is
much more deficient than existing knowledge on the quality of feeds used
in animal nutrition.

Document Number: 539

Wheat transformation technology has progressed rapidly during the past
decade. initially, procedures developed for protoplast isolation and
culture, electroporation- and polyethylene glycol (PEG)-induced DNA
transfer enabled foreign genes to be introduced into wheat cells. The
development of biolistic (microprojectile) bombardment procedures led to
a more efficient approach for direct gene transfer. More recently,
Agrobacterium-mediated gene delivery procedures, initially developed for
the transformation of rice, have also been used to generate transgenic
wheat plants. This review summarises the considerable: progress in wheat
transformation achieved during the last decade.
An increase in food production is essential in order to sustain the
increasing world population. This could be achieved by the development
of higher yielding varieties with improved nutritional quality and
tolerance to biotic and abiotic stresses. Although conventional breeding
will continue to play a major role in increasing crop yield,
laboratory-based techniques, such as genetic transformation to introduce
novel genes into crop plants, will be essential in complementing
existing breeding technologies.
A decade ago, cereals were considered recalcitrant to transformation.
Since then, a significant research effort has been focused on cereals
because of their agronomic status, leading to improved genetic
transformation procedures (Bommineni and Jauhnr 1997). initially, the
genetic transformation of cereals relied on the introduction of DNA into
protoplasts and the subsequent production of callus from which fertile
plants were regenerated. More recently, major advances have been
accomplished in the regeneration of fertile plants from a range of
source tissues, providing an essential foundation for the generation of
transgenic plants. This review summarises procedures, victors and target
tissues used for transformation, highlights the limitations of current
approaches and discusses future trends. The citation of references is
limited, where possible, to the most relevant or recent reports.

Document Number: 540

Flavonoids are common to most vascular plants and are postulated to be
involved in a number of plant functions. Leguminous plants are
intriguing materials for studies on flavonoids for several reasons.
First, legumes produce flavonoids with specific chemical structures i.e.
isoflavonoids and 5-deoxy(iso)flavonoids. Second, leguminous flavonoids
are postulated to play important roles both as defensive compounds
against pathogenic microorganisms and as chemical signals in symbiotic
nitrogen fixation. Third, as constituents of food and forage, leguminous
flavonoids have striking biological activity in humans and animals. This
article reviews the chemical structure, biological activity, and
biosynthesis of leguminous flavonoids. Recent achievements in the
identification and characterization of cytochrome P450 monooxygenases,
which play important roles in (iso) flavonoid biosynthesis, are also
introduced. Perspectives of molecular genetic studies on flavonoids with
the model legume are discussed.

Document Number: 541

Crop biotechnology offers great potentials to contribute to significant
increases in global food production in a sustainable way. Still, the
economic repercussions of biotechnology for developing countries are
largely unknown and have been the subject of acute controversy over the
last few years. This study deals with the topic and provides some
preliminary empirical results. At first, an analytical framework for the
ex ante evaluation of biotechnology products is developed, which is then
used within three case studies:
- banana tissue culture technology in Kenya,
transgenic sweetpotato virus and weevil resistance technology in Kenya
and
- transgenic potato virus resistance technology in Mexico.
The case studies underscore that biotechnology offers efficient
solutions to a wide range of agronomic problems in developing countries,
with concomitant advantages to food producers and consumers.
Biotechnological innovations, which help reduce biotic production
constraints, are suitable for small- and large-scale farmers alike.
Since pest and disease problems are often more severe for smallholders,
the benefit potentials are even greater for this group of farmers.
Especially transgenic crops can easily be integrated into traditional
farming systems because the technology is packaged into the seed and can
be reproduced by the farmers themselves. Apart from the technological
characteristics, however, institutional aspects also influence the
actual socioeconomic outcome. Scenario simulations under different
policy assumptions depict how undesired equity effects could be
prevented.

Document Number: 542

Plants are important resources that have been providing us food from the
earliest times. The rapid advances that have taken place in plant
genetic engineering have made it possible to modify plants to increase
food production and contribute to environmental purification. Transgenic
plants are gaining increasing attention from the industry as a natural
bioreactor for the production of industrial and chemical products.
Useful expression systems based on promoters to optimize transgene
expression in plant cells, hold the key to maximizing the potential of
this concept of molecular-farming or industrial plants. This review,
which is devoted to the use of plants for heterologous protein
production, is divided into three parts. First, we introduce the nature
of plant promoters and strategies for the isolation of novel promoters.
In the second part, various promoters showing high-level constitutive,
organ-specific, or inducible expression, are summarized as useful tools
for realizing the efficient transcription of transgenes. Finally,
problems in the expression of foreign gene in plant cells and future
prospects in plant biotechnology are discussed.

Document Number: 543

Cellulosic materials are the major components of fibrous biomass
produced as a result of photosynthesis and are considered as a reservoir
of solar energy and organic materials. In order to cope with the
problems of food and energy shortages expected in the near future,
biotechnologists are encouraged to develop new technologies for the more
effective utilization of the world's sustainable resources, i.e.,
biomass. One way is to engineer microorganisms and animals with the
capability of digesting and utilizing cellulosic materials, and plants
which can be easily degraded by cellulolytic enzymes. In this article,
we summarize recent studies on the molecular breeding of cellulolytic
organisms for biomass utilization along with some considerations
regarding cellulolytic enzymes.

Document Number: 544

Mycotoxins are fungal secondary compounds that are toxic to vertebrates.
Their presence in food and feeds, as the result of fungal disease in
crops, can present a danger to animal or human health. Many mycotoxins
have also been shown to be phytotoxic and in some cases, such as with
trichothecenes produced by the wheat head blight fungus Fusarium
graminearum, mycotoxins may act as virulence factors.
Antibiotic-producing organisms, including fungi, protect themselves from
their own toxins by metabolic alteration of the compound, modification
of the target site of action or by exporting the compound to the
extracellular space. We have tested the effectiveness of adapting two of
these strategies, metabolic alteration and extracellular transport, to
protect plant cells from the deleterious effects of the trichothecene
4,15-diacetoxyscirpenol (DAS). Tobacco plants were transformed with
either the Saccharomyces cerevisiae gene PDR5, which encodes a
multi-drug transporter, or with the Fusarium sporotrichioides gene
TRI101, which encodes a trichothecene 3-O-acetyltransferase. Both genes
conferred significant increased tolerance to DAS as measured by a
sensitive seed germination assay. Expression of PDR5 or TRI101 in a
seed-specific manner in crop plants such as wheat could lower the
incidence of head blight as well as reduce mycotoxin levels within the
seed. (C) 2000 Elsevier Science Ireland Ltd. 

Document Number: 545

Two recent major drivers of change in the food industry, particulary
with plant-based foods, have been gene technologies and the entry of
intellectual property into many areas of strategic and applied biology
of the principal food plants. It seems likely that for the next few
years advances in food processing will rely on plant breeding which
takes advantages of the increasing knowledge base coming from gene
technologies but which does not rely on the involvement of introduced
transgenes. New diagnostics and assays will allow grains and other foods
to be equipped with changes in their protein, lipid, starch and other
biochemical components so that they better fit food functionalities and
product quality requirements. Subsequently, genetically modified foods
are likely to be accepted as consumers become more familiar with what is
actually involved in bringing about the production of better and safer
foods by these methods. Importantly, many improvements will involve
adjustments without the addition of any new components. These will
assist acceptance of the technologies. It seems inevitable that
transgenic crops will eventually be the norm in our food production
systems because the new technologies promise such significant gains in
efficiencies on farm with corresponding increases in the sustainability
and reliability of the production system. They also will provide for an
unprecedented differentiation of raw materials designed to meet the
demands of the increasingly discriminating consumer.

Document Number: 546

Mercury poisoning has become a problem of current interest as a result
of environmental pollution on a global scale. Natural emissions of
mercury form two-thirds of the input; manmade releases form about
one-third. Considerable amounts of mercury may be added to agricultural
land with sludge, fertilizers, lime, and manures. The most important
sources of contaminating agricultural soil have been the use of organic
mercurials as a seed-coat dressing to prevent fungal diseases in seeds.
In general, the effect of treatment on germination is favorable when
recommended dosages are used. Injury to the seed increases in direct
proportion to increasing rates of application. The availability of soil
mercury to plants is low, and there is a tendency for mercury to
accumulate in roots, indicating that the roots serve as a barrier to
mercury uptake. Mercury concentration in aboveground parts of plants
appears to depend largely on foliar uptake of Hg-0 volatilized from the
soil. Uptake of mercury has been found to be plant specific in
bryophytes, lichens, wetland plants, woody plants, and crop plants.
Factors affecting plant uptake include soil or sediment organic content,
carbon exchange capacity, oxide and carbonate content, redox potential,
formulation used, and total metal content. In general, mercury uptake in
plants could be related to pollution level. With lower levels of mercury
pollution, the amounts in crops are below the permissible levels.
Aquatic plants have shown to be bioaccumulators of mercury. Mercury
concentrations in the plants (stems and leaves) are always greater when
the metal is introduced in organic form. In freshwater aquatic vascular
plants, differences in uptake rate depend on the species of plant,
seasonal growth-rate changes, and the metal ion being absorbed. Some of
the mercury emitted from the source into the atmosphere is absorbed by
plant leaves and migrates to humus through fallen leaves. Mercury-vapor
uptake by leaves of the Cg species oats, barley, and wheat is five times
greater than that by leaves of the C-4 species corn, sorghum, and
crabgrass. Such differential uptake by C-3 and C-4 species is largely
attributable to internal resistance to mercury-vapor binding. Airborne
mercury thus seems to contribute significantly to the mercury content of
crops and thereby to its intake by humans as food. Accumulation,
toxicity response, and mercury distribution differ between plants
exposed through shoots or through roots, even when internal mercury
concentrations in the treated plants are similar. Throughfall and
litterfall play a significant role in the cycling and deposition of
mercury. The possible causal mechanisms of mercury toxicity are changes
in the permeability of the cell membrane, reactions of sulphydryl (-SH)
groups with cations, affinity for reacting with phosphate groups and
active groups of ADP or ATP, and replacement of essential ions, mainly
major cations. In general, inorganic forms are thought to be more
available to plants than are organic ones.
Plants can be exposed to mercurials either by direct administration as
antifungal agents, mainly to crop plants through seed treatment or
foliar spray, or by accident. The end points screened are seed
germination, seedling growth, relative growth of roots and shoots, and,
in some case, studies of leaf-area index, internode development, and
other anatomical characters. Accidental exposures occur through soil,
water, and air pollution. The level of toxicity is usually tested under
laboratory conditions using a wide range of concentrations and different
periods of exposure. Additional parameters include biochemical assays
and genetical studies. The absorption of organic and inorganic mercury
from soil by plants is low, and there is a barrier to mercury
translocation from plant roots to tops. Thus, large increases in mercury
levels in soil produce only modest increases in mercury levels in plants
by direct uptake from soil. Injuries to cereal seeds caused by organic
mercurials has been characterized by abnormal germination and
hypertrophy of the roots and coleoptile.
Mercury affects both light and dark reactions of photosynthesis.
Substitution of the central atom of chlorophyll, magnesium, by mercury
in vivo prevents photosynthetic light harvesting in the affected
chlorophyll molecules, resulting in a breakdown of photosynthesis. The
reaction varies with light intensity. A concentration and time-dependent
protective effect of GSH seems to be mediated by the restricted uptake
of the metal involving cytoplasmic protein synthesis. Plant cells
contain aquaporins, proteins that facilitate the transport of water, in
the vacuolar membrane (tonoplast) and the plasma membrane. Many
aquaporins are mercury sensitive, and in AQP1 a mercury-sensitive
cysteine residue (Cys-189) is present adjacent to a conserved
Asn-Pro-Ala motif. At low concentrations mercury has a toxic effect on
the degrading capabilities of microorganisms. Sensitivity to the metal
can be enhanced by a reduction in pH, and tolerance of mercury by
microorganisms has been found to be in the order: total population >
nitrogen fixers > nitrifiers. Numerous experiments have been carried out
to study the genetic effects of mercury compounds in experimental test
systems using a variety of genetic endpoints. The most noticeable and
consistent effect is the induction of c-mitosis through disturbance of
the spindle activity, resulting in the formation of polyploid and
aneuploid cells and c-tumors. Organomercurials have been reported to be
200 times more potent than inorganic mercury. Exposure to inorganic
mercury reduces mitotic index in the root-tip cells and increases the
frequency of chromosomal aberrations in degrees directly proportional to
the concentrations used and to the duration of exposure. The period of
recovery after removal of mercury is inversely related to the
concentration and duration of exposure.
Bacterial plasmids encode resistance systems for toxic metal ions,
including Hg2+, functioning by energy-dependent efflux of toxic ions
through ATPases and chemiosmotic cation-proton antiporters. The
inducible mercury resistance (mer) operon encodes both a mercuric ion
uptake and detoxification enzymes. In gram-negative bacteria a
periplasmic protein, MerP, an inner-membrane transport protein, MerT,
and a cytoplasmic enzyme, mercuric reductase, the MerA protein, are
responsible for the transport of mercuric ions into cells and their
reduction to elemental mercury, Hg(II). In Thiobacillus ferrooxidans, an
acidophilic chemoautotrophic bacterium sensitive to mercury ions, a
group of mercury-resistant strains, which volatilize mercury, has been
isolated. The entire coding sequence of the mercury-ion resistance gene
has been located in a 2.3 kb fragment of chromosomal DNA (encoding
56,000 and 16,000 molecular-weight proteins) from strain E-15 of
Escherichia coli. Higher plants and Schizosaccharomyces pombe respond to
heavy-metal stress of mercury by synthesizing phytochelatins (PCs) that
act as chelators. The strength of Hg(II) binding to glutathione and
phytochelatins follows the order: gamma
Glu-Cys-Gly<(<gamma>Glu-Cys)(2)Gly<(<gamma>Glu-Cys)(3)Gly<(<gamma>Glu-Cy
s)(4)Gly. Suspension cultures of haploid tobacco, Nicotiana tabacum,
cells were subjected to ethyl methane sulfonate to raise
mercury-tolerant plantlets. HgCl2-tolerant variants were selected from
nitrosoguanidine (NTG)-treated suspension cell cultures of cow pea,
Vigna unguiculata, initiated from hypocotyl callus and incubated with 18
mug/ml HgCl2. Experiments have been carried out to develop
mercury-tolerant plants of Hordeum vulgare through previous exposure to
low doses of mercury and subsequent planting of the next generation in
mercury-contaminated soil. Phytoremediation involves the use of plants
to extract, detoxify, and/or sequester environmental pollutants from
soil and water. Transgenic plants cleave mercury ions from methylmercury
complexes, reduce mercury ions to the metallic form, take up metallic
mercury through their toots, and evolve less toxic elemental mercury.
Genetically engineered plants contain modified forms of bacterial genes
that break down methyl mercury and reduce mercury ions. The first gene
successfully inserted into plants was merA, which codes for a mercuric
ion reductase enzyme, reducing ionic mercury to the less toxic elemental
form. MerB codes for an organomercurial lyase protein that cleaves
mercury ions from highly toxic methyl mercury compounds. Plants with the
merB gene have been shown to detoxify methyl mercury in soil and water.
Both genes have been successfully expressed in Arabidopsis thaliana,
Brassica (mustard), Nicotiana tabacum (tobacco), and Liriodendron
tulipifera (tulip poplar). Plants currently being transformed include
cattails, wild rice, and Spartina, another wetland plant. The problem of
mercury contamination can be reduced appreciably by combining the
standard methods of phytoremediation-removal of mercury from polluted
areas through scavenger plants-with raising such plants both by routine
mutagenesis and by genetic engineering. The different transgenics raised
utilizing the two genes merA and merB are very hopeful prospects.

Document Number: 547

A Research Committee was established by the Weed Science Society of
America to outline the direction of weed science research during the
next decade. Weeds adversely affect humans in both agricultural and
nonagricultural environments. It is the opinion of the research
committee that weed science will be advantageously positioned for the
future if research focuses on research decision processes, weed biology
and ecology, weed control and management practices, herbicide
resistance, issues related to transgenic plants, environmental issues,
and potential benefits of weeds. These future weed science research
directions endorse those of the commodity and grower input group
Coalition for Research on Plant Systems (CROPS)'99, a U.S. Department of
Agriculture (USDA)-supported initiative. The future of weed science is
dependent on a, joint effort from industry, government regulators, and
the public sector consisting of grower groups, as well as USDA,
Agriculture and Agri-Food Canada (AAFC), and university researchers. It
is our opinion that efforts spent on these research areas will benefit
not only growers, commodity groups, homeowners, and industry, but
society at large, through the maintenance and improvement of the food
and fiber production system, and the environment in North America.

Document Number: 548

Recent advancements in agricultural biotechnology have created a need
for analytical techniques to determine introduced proteins in crops
enhanced through modern biotechnology techniques. These proteins are
expressed in plant tissues and may be present in food ingredients.
Immunoassays are ideally suited for protein detection and may be used as
both quantitative and threshold methods. Microplate ELISA and lateral
flow devices are two of the most commonly used immunoassay formats for
agricultural biotechnology applications. This paper provides general
background information and a discussion of criteria for the validation
and application of immunochemical methods to the analysis of proteins
introduced into plants and food ingredients using biotechnology methods.
It is the result of a collaborative effort of members of the Analytical
Environmental Immunochemical Consortium. This collaborative effort
represents the combined expertise of several organizations to reach
consensus on establishing guidelines for the validation and use of
immunoassays. Further, the paper offers developers and users a
consistent approach to adopting the technology as well as aid in
producing accurate and meaningful results.

Document Number: 549

This paper reviews the possibility and limits for increasing the content
and bioavailability of iron (Fe), zinc (Zn) and calcium (Ca) in edible
parts of staple crops, such as cereals, pulses, roots and tubers as a
way to combat mineral deficiencies in human populations. Theoretically,
this could be achieved by increasing the total level of Fe, Zn and Ca in
the plant foods, while at the same time increasing the concentration of
compounds which promote their uptake (ascorbic acid), and/or by
decreasing the concentration of compounds which inhibit their absorption
(phytic acid or phenolic compounds). The content of Zn and Ca in grains
and fruits can in some cases be increased through soil and/or foliar
applications of Zn and Ca fertilisers. Plant breeding and genetic
engineering techniques, however, have the greatest potential to increase
Fe and Zn content in grains, roots and tubers. The possibility of
enhancing Ca and ascorbic acid content in plant foods by plant breeding
and genetic engineering remained to be explored. The critical factor is
to ensure that the extra minerals have an adequate bioavailability for
man. Given the important role of phytic acid and polyphenols in plant
physiology, reducing the levels of these compounds in the edible parts
of plants does not appear to be wise although introduction of phytases
which are active during digestion is an exciting possibility. (C) 2000
Society of Chemical Industry.

Document Number: 550

There is growing recognition that many phenolic secondary metabolites
present in foodstuffs may possibly exert beneficial effects on human
health. This may to some degree be mediated via antioxidant actions, but
a range of more specific pharmacological effects have also been
proposed. Given this background, there may be favourable consequences
for the general health of Western populations as a result of optimising
the phenolic content of the diet. This paper reviews what is known of
the function of phenolics both in the plant and in man. It also
describes current understanding of the biosynthesis of phenolics in
plants, with emphasis on where potential controlling steps may exist.
Finally, advances in identification and isolation of the genes coding
for phenolic biosynthetic enzymes or regulatory proteins are also
summarised. Taken together, this information provides a basis for
attempts to modify and optimise the phenolic content of food crops,
using either conventional plant breeding along with manipulation of
agronomic practices, or else the more targeted approaches of modern
molecular biology. (C) 2000 Society of Chemical Industry.

Document Number: 551

The presence of genetically modified organisms (GMOs) in food products
is usually ascertained by the polymerase chain reaction (PCR) or nested
PCR if sensitivity has to be increased. Since most, if not all, GMO
products are of plant origin, the target sequences are the 35S promoter
or NOS terminator. The extreme sensitivity of nested PCR can be
misleading if the results are not interpreted correctly, since
contamination of non-GMO products with residual amounts of GMO may be
positive, We report that breaded turkey breast cutlets labeled as
containing GMO soybean products were actually prepared from wheat flour
that had been contaminated with transgenic soybean.

Document Number: 552

The effect of fructans in human nutrition became more and more important
during the last years and in the same way the knowledge about the
fructan metabolism in plants and the production of fructans from plants.
About 15% of higher plants and also numerous crops store fructans. The
use of fructans is either indirectly as food ingredients or directly by
the use of vegetables. Fructans are mostly gained from chicory, but the
use of Jerusalem artichoke, transgenic potato and sugar beet lines are
also discussed. Vegetable crops like onion, leek, asparagus, and sprouts
from cereals contain rather high amounts of fructans and may become more
important in the near future, e.g. as "functional food".
Many foods can be enriched with fructans as dietary fibers without any
negative impact on the taste of the product, With an additional uptake
of 10-15 g of fructans per day, the recommended daily uptake of dietary
fibers (30 g) could be reached. Fructans are stimulating the growth of
specific microorganisms in the colon (e.g. Bifidobacteria) with a
general positive health effect.

Document Number: 553

Plants and most animals can synthesize their own L-ascorbic acid
(vitamin C), but a mutation in the L-gulono-gamma-lactone oxidase gene
in the primate lineage makes it necessary for humans to acquire this
vital compound from their diet. Despite the fact that plants and animals
synthesize ascorbic acid via different pathways, transgenic tobacco and
lettuce plants expressing a rat cDNA encoding L-gulono-gamma-lactone
oxidase accumulated up to seven times more ascorbic acid than
untransformed plants. These results demonstrate that basal levels of
ascorbic acid in plants can be significantly increased by expressing a
single gene from the animal pathway.

Document Number: 554

With increasing population and decreasing area of land available for
food production the development and use of genetically modified (GM)
crops is considered as an important tool to ensure global food security.
The rigorous safety evaluation, which can take up to 10 years by
independent Regulatory Agencies prior to final approval of GM crops, is
noted. The rapid uptake of GM technology is illustrated by the increase,
between 1996/1999, from 2 to 40 million hectares and that while 72% is
grown in the USA a further 15 countries grew GM crops. The paper
discusses a number of safety issues including antibiotic resistance
marker genes, potential allergenicity, substantial equivalence,
consumption of transgenic protein and transgenic DNA and their detection
in milk, meal and eggs. Far from increasing the risk of allergenicity GM
technology can offer the opportunity to reduce/eliminate protein
allergens that occur naturally in foods such as rice, wheat and peanuts
and that GM crops are compositionally (laboratory analysis) and
nutritionally (animal feeding studies) equivalent to their conventional
counterparts. The paper notes that while concern has been expressed
about the possible accumulation of transgenic protein and DNA in milk,
meal and eggs, and the possible implications of this on animal and human
health, the regulatory agencies and the World Health Organisation
concluded that there is no inherent risk in consuming DNA, including
that fi om GM crops as mammals have always consumed significant
quantities of DNA from a wide variety of sources, including plants,
animals, bacteria, parasites and viruses. To date transgenic protein and
DNA have not been found in milk, meat and eggs. The paper concludes, by
providing numerous examples of the potential benefits to the farmer,
consumer and the environment in both developed and developing countries,
with particular emphasis on the ability of GM technology to increase
crop yield, improve food quality and to reduce the environmental impact
of agriculture by amongst other things the lower use of less toxic
agrochemicals.

Document Number: 555

Transgenic foods are a favourable application of biotechnology in the
agroalimentary area.
Directives, regulations, and recommendations both comunitary and
national have been issued to regulate the emission in the environment of
genetically modified organisms and the immission bn market of foods and
ingredients genetically modified.
Some proposals about labelling, formal checks, genetic markers and
minimal limit of validity are still to be completely determined.
Further technical and scientific contributions are necessary to grant
the health of genetically modified food consumers, also for possible
longterm effects.

Document Number: 556

Methods for the quantification of transgenic insect-resistant Bt maize
by single- and dual-competitive PCR were developed. The analysis of
mixtures of DNA solutions, as well as of maize flours containing defined
amounts of Bt maize, demonstrated the usefulness of single-competitive
PCR based on coamplification of the CDPK promoter/cry1A(b) gene region
of Bt maize and an internal standard. Upon heat treatment of DNA
solutions and maize flour, respectively, the recovery of the Bt
proportion compared to the starting material determined by
single-competitive PCR decreased significantly. This systematic error
could be compensated for by using a dual-competitive approach based on
PCR quantification of the transgenic target sequence of Bt maize and of
the maize specific invertase gene (ivr1).

Document Number: 557

Today DNA-based techniques are very common for the detection of
genetically modified organisms (GMOs) in food products. For fast and
easy detection of GMOs, polymerase chain reaction (PCR) screening
methods, which amplify common transgenic elements, are applied in
routine analysis. These techniques do not allow differentiation between
GMOs and the natural occurrence of transgenic elements, such as the
35S-promoter of cauliflower mosaic virus (CaMV) or the NOS-terminator of
Agrobacterium tumefaciens, and thus may result in false-positive
detection of GMOs. In this study we evaluated three different existing
35S screening systems and report the development of two new
CaMV-specific PCR systems. These PCR systems based on CaMV-specific
genes allow the identification of positively screened 35S food samples
as naturally virus-infected products or plants. Seven food samples
tested positive in routine 35S screening analysis and negative in GMO
specific systems were investigated using the new virus-specific PCR
systems. In all seven samples CaMV was detected.

Document Number: 558

Despite the application of 2.5 million tons of pesticides worldwide,
more than 40% of all potential food production is lost to insect, weed,
and plant pathogen pests prior to harvest. After harvest, an additional
20% of food is lost to another group of pests. The use of pesticides for
pest control results in an estimated 26 million human poisonings, with
220,000 fatalities, annually worldwide. In the United States, the
environmental and public health costs for the recommended use of
pesticides total approximately $9 billion/yr. Thus, there is a need for
alternative non-chemical pest controls, and genetic engineering
(biotechnology) might help with this need. Disease and insect pest
resistance to various pests has been slowly bred into crops for the past
12,000 years; current techniques in biotechnology now offer
opportunities to further and more rapidly improve the non-chemical
control of disease and insect pests of crops. However, relying on a
single factor, like the Bacillus thuringiensis toxin that has been
inserted into corn and a few other crops for insect control, leads to
various environmental problems, including insect resistance and, in some
cases, a threat to beneficial biological control insects and endangered
insect species. A major environmental and economic cost associated with
generic engineering applications in agriculture relates to the use of
herbicide resistant crops (HRC). In general, HRC technology results in
increased herbicide use but no increase in crop yields. The heavy use of
herbicides in HRC technology pollutes the environment and can lead to
weed control costs for farmers that may be 2-fold greater than standard
weed control costs. Therefore, pest control with both pesticides and
biotechnology can be improved for effective, safe, economical pest
control.

Document Number: 559

Parenteral and oral administration of autoantigens can induce immune
tolerance in autoimmune diseases. Prophylactic therapy based on oral
administration of human autoantigens is not, however, feasible when
sufficient quantities of candidate autoantigens are not available.
Transgenic plants that express high levels of recombinant proteins would
allow large quantities of autoantigens to be produced at relatively low
costs. In addition, transgenic food would provide a simple and direct
method of delivering autoantigens. The production and the
characterization of transgenic tobacco and carrot plants expressing
human GAD65, a major autoantigen in human insulin-dependent diabetes
mellitus (IDDM), is reported. Immunogold labeling and electron
microscopy of transgenic tobacco tissue shows the selective targeting of
human GAD65 to chloroplast tylacoids and mitochondria. In planta
expressed GAD65 has a correct immunoreactivity with IDDM-associated
autoantibodies and retains enzymatic activity, a finding that suggests a
correct protein folding. In transgenic tobacco and carrot the expression
levels of human GAD65 varies between 0.01% and 0.04% of total soluble
proteins. Transgenic edible plant organs are now available to study the
feasibility of inducing immune tolerance in IDDM animals by oral
administration of GAD65.

Document Number: 560

The detection of genetically modified foodstuffs is becoming both a food
sales and legal necessity. This study reports a rapid DNA
extraction/PCR-based method for the detection of genetically modified
soya (GMS) and maize (GMM) in mixed samples of transgenic and unmodified
soybeans and maize kernels, and a variety of processed samples including
soya flour, soya protein isolates, extruded defatted soya, acid- and
alcohol-precipitated soya concentrates, soya lecithin, maize grits,
seasoned corn puffs and salted corn chips. The presence of GMS DNA was
determined with two pairs of primers directed towards different GMS
target sequences and GMM by one primer pair. In addition, a multiplex
PCR reaction which utilises an internal positive control was developed
for both genetically modified organisms (GMOs). Results indicated that
the methods are sensitive and specific enough to detect GMS down to a
level of 0.01% dry weight in single-product PCRs and 0.1% in multiplex
PCRs and GMM down to 0.001% dry weight in single-product PCRs and 0.01%
in multiplex PCR. The methods are considered to represent a viable route
for the commercial detection of GMS and GMM in foodstuffs.

Document Number: 561

A fast and quantitative method was developed to detect transgenic
"Maximizer" maize "event 176" (Novartis) and "Roundup Ready" soybean
(Monsanto) in food by real-time quantitative PCR. The use of the ABI
Prism 7700 sequence detection system allowed the determination of the
amplified product accumulation through a fluorogenic probe (TaqMan).
Fluorescent dyes were chosen in such a way as to coamplify total and
transgenic DNA in the same tube. Using real-time quantitative PCR, 2 pg
of transgenic or total DNA per gram of starting sample was detected in 3
h after DNA extraction and the relative amounts of "Maximizer" maize and
"Roundup Ready" soybean in some representative food products were
quantified.

Document Number: 562

Analytical methods for authenticity testing have been described for all
types of food and can give us important indications for analytical
strategies to be developed for the detection and quantitation of
genetically modified foods. Transgenic plants contain newly introduced
traits or marker genes that are expressed and should be detectable by
DNA or protein-based methods. Recent literature clearly favours PCR as
the method of choice for the identification of CMO-derived food.
Quantitative competitive PCR is an important extension of this method
allowing to control labelling and maximum limits to be set for
genetically modified crops in non-GMO-crops. (C) 1999 Elsevier Science
Ltd. 

Document Number: 563

Two different PCR-based approaches for the quantitative analysis of
genetically modified organism (GMO) - components in foods are presented
using Soybean derived samples as an example. The first method - a double
competitive PCR - is well suited to determine threshold levels of GMO
content in food,The other - PCR on-line measurement - is suited to
determine ratios of transgenic versus non-transgenic component. Both
methods provide a means to alleviate the problems of standardisation
encountered with simple qualitative PCR approaches and will allow to
cope with threshold levels for GMO, once issued by legislative bodies.
(C) 1999 . 

Document Number: 564

The principle of direct detection of recombinant DNA in food by the
polymerase chain reaction (PCR) is discussed following the three main
steps: DNA-extraction, amplification by PCR and verification of PCR
products.
Suitable methods for genomic DNA isolation from homogenous,
heterogeneous, low DNA containing matrices (e.g. lecithin), gelatinising
material (e.g. starch), derivatives and finished products based on
classical protocols and/or a combination with commercially available
extraction kits are discussed. Various factors contribute to the
degradation of DNA such as hydrolysis due to prolonged heat-treatment,
nuclease activity and increased depurination and hydrolysis at low pH.
The term "DNA quality" is defined as the degree of degradation of DNA
(fragment size less than 400 bp in highly processed food) and by the
presence or absence of potent inhibitors of the PCR and is, therefore, a
key criterion. In general, no DNA is detectable in highly heat-treated
food products, hydrolysed plant proteins (e.g. soya sauce), purified
lecithin, starch derivatives (e.g. maltodextrins, glucose syrup) and
defined chemical substances such as refined soya oil.
If the nucleotide sequence of a target gene or stretch of transgenic DNA
is already known specific primers can be synthesised and the segment of
rDNA amplified. Detection limits are in the range 20 pg-10 ng target DNA
and 0.0001-1% mass fraction of GMO. Amplification products are then
separated by agarose gel electrophoresis and the expected fragment size
estimated by comparison with a DNA molecular weight marker.
Several methods are used to verify PCR results and they vary in
reliability, precision and cost. They include specific cleavage of the
amplification products by restriction endonucleases or the more
time-consuming, but also more specific, transfer of separated
PCR-products onto membranes (Southern Blot) followed by hybridisation
with a DNA probe specific for the target sequence. Alternatively, PCR
products may be verified by direct sequencing. Nested-PCR assays
combines high specificity and sensitivity.
Methods for the screening of 35S-promoter, NOS;terminator and other
marker genes used in a wide range of GMOs, the specific detection of
approved products such as FlavrSavr(TM) tomatoes, Roundup Ready(TM)
Soya, Bt-maize 176 and official validated methods for potatoes and
genetically modified micro-organisms, that have a model character, are
available. Methods to analyse new GMO products are being validated by
interlaboratory tests and new techniques are in development (e,g. EC
project: DMIF-GEN). However, these efforts may be hampered by the lack
of availability of GMO reference material as well as specific sequence
information which so far can only be obtained from the suppliers. (C)
1999 . 

Document Number: 565

Transgenic animals are useful tools for the study of biological
fractions of proteins and secondary gene products synthesized by the
action of protein catalysts. Research in nutrition and allied fields is
benefiting from their use as models to contrast normal and altered
metabolism. Although food, nutritional products, and ingredients from
transgenic animals have not yet reached consumers, the technologies for
their production are maturing and yielding exciting results in
experimental and farm animals. Regulatory governmental bodies are
already issuing guidelines and legislation in anticipation of the advent
of these products and ingredients. This review summarizes available
technology for the production of transgenic animals, discusses their
scientific and commercial potential, and examines ancillary issues
relevant to the field of nutrition. (C) Elsevier Science Inc. 1999. All
rights reserved.

Document Number: 566

According to FAO, China has a population of 133 million sheep, 171
million goats and 300 thousand antler deer. The population of goats is
the largest in the world and of sheep the second largest. The total
production of mutton is 2.4 million ton, which represents only 4.1% of
the total meat production of 59.2 million ton in China. The most
important strategy of China is to improve the breeding of sheep and
goats, and increase their meat output by utilizing forage and grassland
resources efficiently. Acid produce enough protein food for the
increasing population through the use of new biotechnology. Research in
biotechnology of small ruminants did not begin in China until 1970, but
there has been much progress in recent years. The techniques of ovine
and caprine embryo transfer, embryo freezing, embryo splitting, sex
determination, in vitro, and caprine nuclear transfer has all been
successful. The application of artificial insemination in antler deer
has just begun and in 1996, investigations in embryo transfer have also
begun. From 1995 to 1996, the rate of increase of ovine and caprine was
4.3% and 14.1%, respectively, and biotechnology of sheep and goats will
be applied more widely. Research has been conducted on transgenic
animals, improvement of breeding and enhancing their productivity
through biotechnology. Native sheep and goat breeds will be protected
because of the value of their polymorphisms. (C) 1999 Published by
Elsevier Science B.V. 

Document Number: 567

Transgenic tobacco plants exhibiting exogenous cellulase activity were
produced by introduction of a truncated cellulase (t-EgI) gene from
Ruminococcus albus. They accumulated about 0.1-0.5% t-EgI of the total
soluble proteins in the cytoplasm. They were normal in morphology and
growth. t-EgI expressed in the transgenic plants degraded their own cell
wall materials after the cells were disrupted. The plants expressing the
t-egI gene could be used as "cellulase-encapsulating plants" for
livestock and food industries.

Document Number: 568

Little is known about the effect of transgenic plants containing
commonly used marker genes, such as aph(3')II (nptII encoding
neomycinphosphotransferase) and uidA (gus encoding beta-glucuronidase)
on insect feeding behaviour. We report here, for the first time, that
transgenic potato plants containing only nptII and gus marker genes
enhance foliage consumption by the Colorado potato beetle (CPB,
Leptinotarsa decemlineata S.). Transformation of potato cultivar Desiree
was performed with Agrobacterium tumefaciens. Internode explants were
inoculated with different strains of bacteria, carrying either nptII-gus
or nptII alone. A total of 180 transgenic and untransformed control
plants were grown in the greenhouse for the analysis of food consumption
by CPB. For each transformed and untransformed line tested, four
bioassays were conducted each consisting of 10 second-instar larvae
feeding independently on a 2 cm diameter leaf disc for 20 h. Our data
show up to 50% increase of mean foliage consumption on plants
transformed with the nptII-gus construct, indicating that transgenic
plants containing these marker genes can affect the feeding behaviour of
the insects. These results were obtained from the 'primary regenerants'
(R-0 lines) as well as from tuber-derived plants (R-1 lines). Further
tests with transgenic plants containing the nptII marker gene only,
showed no significant difference in feeding when compared to
untransformed control plants, allowing us to rule out a direct effect of
this marker gene on foliage consumption by the insect larvae. It is
suggested that gus protein is involved in the increase of foliage
consumption by CPB.

Document Number: 569

Soybean (Glycine max) is the most important oil crop worldwide. Because
of its high content of oil and protein there are many uses in the food
and non-food sector. The cultivated soybean originates from the wild
species Glycine soja. Numerous other wild Glycine species form a huge
reservoir of genetic resources that are slowly being utilized by
breeding. Mutagenic and genetic engineering studies of the soybean
genome have resulted in drastic alterations of fatty and amino acid
composition. The introduction of a bacterial gene for tolerance to the
herbicide glyphosate into the soy-bean genome has led to transgenic
lines which are commercially grown on a large scale.

Document Number: 570

The high-molecular-weight (HMW) subunits of wheat glutenin are the major
determinants of the gluten visco-elasticity that allows wheat doughs to
be used to make bread, pasta and other food products. In order to
increase the proportions of the HMW subunits, and hence improve
breadmaking performance, particle bombardment was used to transform
tritordeum, a fertile amphiploid between wild barley and pasta wheat,
with genes encoding two HMW glutenin subunits (1Ax1 and 1Dx5). Of the 13
independent transgenic lines recovered (a transformation frequency of
1.4%) six express the novel HMW subunits at levels similar to, or higher
than, those of the endogenous subunits encoded on chromosome 1B.
Small-scale mixograph analysis of T(2) seeds from a line expressing the
transgene for 1Dx5 indicated that the addition of novel HMW subunits can
result in significant improvements in dough strength and stability, thus
demonstrating that transformation can be used to modify the functional
properties of tritordeum for improved breadmaking.

Document Number: 571

Genetically modified, i.e. transgenic crops, are now being grown on
several million acres throughout the world, mainly in North America.
Most of these 'first generation' products have specific agronomic traits
designed to improve the efficiency of production. For example herbicide
tolerant soybean and insect resistant corn are the two most widely grown
transgenic crops. Increasingly, the new transgenic varieties under test
contain product quality or other higher value traits intended to provide
specific benefits to the end user whether it be the producer of
specialist chemicals or the consumer. This review describes examples of
these 'second generation' traits and attempts to predict the range of
household, medical, industrial and environmental products that might
become available over the next 20 years. (C) 1999 Annals of Botany
Company.

Document Number: 572

The compositions of transgenic potatoes with intact and modified genes
for soybean glycinin were compared with those of control potatoes;
non-transgenic or transgenic ones with at vector. The expression levels
of glycinin proteins in the transgenic potato tubers were approximately
12-31 mg g(-1)-total soluble protein. With reference to six major
components, moisture, protein, lipid, fiber, ash and carbohydrate, there
were no significant differences between transgenic and control potatoes.
The levels of glycoalkaloids in transgenic potato tubers, as well as in
transgenic controls, increased in comparison with those in
non-transgenic ones, though the level was within the permitted limit.
The modified glycinin expressed in the transgenic potato tuber was
digested under the simulated gastric conditions. From these results, the
transgenic potatoes with intact and modified glycinin genes are
considered to be as safety utilization for food as non-transgenic
potatoes. (C) 1999 Society of Chemical Industry.

Document Number: 573

A beta-glucuronidase (GUS) gene fusion with the promoter from the potato
Lhca3.St.1 gene, encoding a light-harvesting complex protein, shows
expression in pollen of transgenic tobacco plants. This was established
by histochemical and fluorometric assays, and confirmed by RT-PCR,
Western analysis and GUS activity following gel electrophoresis, The
activity is minimal in early pollen development and high in mature
pollen at anthesis, It shows gametophytic segregation in pollen grains.
Pollen from tobacco plants hemizygous for a single Lhca3.St.1-GUS locus
exhibited 1:1 segregation ratios, whereas dihybrids segregated in the
expected 3:1 ratio. A sequential combination of histochemical staining
for GUS activity and Alexander's stain for pollen viability, offers a
convenient system to eliminate artefacts in the histochemical staining,
identify inviable pollen and accurately score pollen segregation. The
dual staining approach also confirmed gametophytic segregation in
tobacco plants hemizygous for the GUS gene under transcriptional control
of a doubled CaMV 35S promoter. Gametophytic segregation establishes
that the GUS activity is determined by the genetic status of the pollen
rather than of the parent plant, and indicates de novo GUS gene
expression and translation during pollen maturation. The unanticipated
activity of the Lhca3.St.1 promoter in pollen has implications for the
biosafety assessment of transgenic plants intended for agricultural use.
Even when a promoter is not generally considered to be active in pollen,
its expression in pollen should be evaluated if effects on pollinating
insects or food safety concerns for products such as honey can be
anticipated.

Document Number: 574

Hexaploid tritordeum is the fertile amphiploid (2n = 6x = 42,
AABBH(ch)H(ch)) between Hordeum chilense and durum wheat. There is huge
generic variability available for breeding this new species obtained
after the synthesis of hundreds of different amphiploids.
Morphologically and agronomically, tritordeum is similar to wheat.
Tritordeums exhibit an uptake nitrogen system with a higher affinity for
nitrate than their wheat parent. Similarly. tritordeum has a higher
water use efficiency than wheat. The rheological and baking
characteristics of hexaploid tritordeum flour are also quite similar to
those of bread wheat, indicating that the H(ch) genome can at least
partially substitute for the D genome. The quality of tritordeum flour
is determined by both the wheat and the barley seed storage proteins.
The H. chilense seed storage proteins show a high degree of variation,
which could be useful as,an additional source of generic variability for
wheat processing (pasta and bread) quality by means of using tritordeum
as the bridge species. Similarly, the tritordeum endosperm protein
composition has been modified through transformation and by chromosome
manipulation with glutenin genes from wheat. (C) 1999 Academic Press.

Document Number: 575

Biotechnology is a key technology in the generation of an adequate food
supply for the ever growing human population. Genetics and reproduction
as fundamental functional prerequisites for successful livestock
production have been important subjects of biotechnological research in
animal science for many decades. Biotechnological procedures developed
so far are to be applied for increasing breeding efficiency in farm
animals, for the preservation of animal genetic resources, for the
improvement of product quality or for new production strategies and
novel animal products. Many of these opportunities are already routine
in animal husbandry, others will be employed according to needs,
suitability and their public perception. The latter has to be addressed
by observing the ethical principles for the application of biotechnology
in animals in selected areas of livestock production in which immediate
benefits are to be obtained. This paper deals with some of the future
perspectives and visions for animal biotechnology in research and field
application. It focusses on cryopreservation of germ cells and embryos,
ovum pickup from ovaries in situ and IVMFC, gender preselection,
cloning, genome analysis and gene mapping, and gene transfer. (C) 1999
Elsevier Science B.V. 

Document Number: 576

Rapeseed is one of the most important oilproducing plants in Germany,
Asia and North America. Rapeseed oil is e.g. used for the production of
salad oil and edible fat. Microscopical analysis of rapeseed pollen is
an important item for the determination of the authencity of honey,
Although the analysis of rapeseed DNA would be therefore of great
interest, Glufosinate resistant rapeseed is one of the prominent
genetically modified plants for cultivation and commercialization in
Europe. The same transgenic sequence is also present in genetically
modified maize and sugar-beets, For surveillance the Novel Food
legislation as well as the labelling of genetically modified seeds and
feeds an appropriate method for the detection of transgenic DNA is
necessary, This paper describes a method for the extraction of DNA from
rapeseed pollen in honey and the specific detection of the new trait
(Glufosinate resistance) in genetically modified rapeseed, maize and
sugar-beets.

Document Number: 577

Foods produced by means of genetic engineering have to be labeled
according to the European Union Regulation No 258/97 (Novel Food
Regulation). Products derived from genetically modified tomatoes
developed by Zeneca are Novel Foods. In order to enable the authorized
food control agencies to supervise compliance with the labeling
requirements a method was developed to identify the transgenic tomatoes.
It is based on the combination of the classical CTAB protocol and a
subsequent nucleic acid purification step with the QIAquick PCR
Purification Kit (Qiagen). Various processed tomato products available
on the Berlin market were investigated applying this method.

Document Number: 578

One of the criterions for labelling of genetically modified foods to the
Novel Food legislation is the presence of transgenic DNA. Therefor
analysis is mostly based on the detection of the transgenic DNA. No
labelling is required for unavoidable residues below a limiting level.
Appropriate methods for the quantification of such residues are therefor
necessary. This paper describes a competitive PCR method for the
quantification of the genetically modified soy beans.

Document Number: 579

The composition of nutritionally and physiologically important molecules
in transgenic rice with the soybean glycinin gene was determined and
compared with that of a nontransgenic control. Except for the levels of
protein, amino acids and moisture, no marked differences were found
between the two kinds of rice. The protein content of the transgenic
rice was about 20% higher than the control(control, 6.5 g/100 g;
transgenic, 8.0 g/100 g) with a concomitantly lower moisture content.
This increased protein content mainly resulted from the increased
glycinin expressed in the transgenic rice, and the protein was
susceptible to gastric and intestinal digestion juices. In parallel with
the increased protein content, some important amino acids lacking in
quantity in normal rice were replenished.

Document Number: 580

Transgenic crops genetically engineered for enhanced insect resistance
should be compatible with other components of IPM for the pest
resistance to be durable and effective. An experimental potato line was
genetically engineered to express an anti-aphid plant protein (snowdrop
lectin, GNA), and assessed for possible interactions of the insect
resistance gene with a beneficial pest predator These extended
laboratory studies are the first to demonstrate adverse tri-trophic
interactions involving a lectin-expressing transgenic crop, a target
pest aphid and a beneficial aphidophagous predator. When adult 2-spot
ladybirds (Adalia bipunctata [L.]) were fed for 12 days on peach-potato
aphids (Myzus persicae Sulzer) colonising transgenic potatoes expressing
GNA in leaves, ladybird fecundity, egg viability and longevity
significantly decreased over the following 2-3 weeks. No acute toxicity
due to the transgenic plants was observed, although female ladybird
longevity was reduced by up to 51%. Adverse effects on ladybird
reproduction, caused by eating peach-potato aphids from transgenic
potatoes, were reversed after switching ladybirds to feeding on pea
aphids from non-transgenic bean plants. These results demonstrate that
expression of a lectin gene for insect resistance in a transgenic potato
line can cause adverse effects to a predatory ladybird via aphids in its
food chain. The significance of these potential ecological risks under
field conditions need to be further evaluated.

Document Number: 581

The aim of the study was to settle a concent for the standardised
description of the potency of a PCR setup with respect to the detection
limit. The main terms used are the theoretical detection limit (L-theor)
and the practical detection limit (L-prac). These detection limits were
determined for the example of two current PCR setups for the detection
of genetically modified soybeans and maize. The Ltheor for the detection
of Roundup Ready soybean DNA with the PCR setup applied was 0.005%
genetically modified organism (GMO)/non-GMO (w/w) (corresponding to 30
copies of the GMO soya genome per single PCR reaction). In pre-mixed
powder preparations (certified reference materials, CRMs) of soya it was
possible to detect 0.1% GMO/non-GMO (w/w), i.e. this was the L-prac. The
Ltheor for Bt maize was 0.005% GMO/non-GMO (w/w), that corresponded to 9
calculated genome copies per single reaction. Without background DNA, as
few as 2 genome copies (0.01 ng GMO DNA) were clearly detectable. As for
soybeans, the L-prac of the available CRMs from transgenic and
conventional maize was 0.1% GMO/non-GMO (w/w). Quantitative systems for
the investigation of foodstuffs are still not available as official
methods, and are still being developed. Determining L-theor and L-prac
values could be important for setting limits for the GMO contents of
foods, when deciding whether they should be labelled. Concerning
labelling limits, it is obvious that the PCR setups investigated have
more sensitive detection thresholds than the limits under discussion for
the labelling of foodstuffs as "genetically modified".

Document Number: 582

Vegetable oils are a major component of human diets, comprising as much
as 25% of average caloric intake. Until recently, it was not possible to
exert significant control over the chemical composition of vegetable
oils derived from different plants. However, the advent of genetic
engineering has provided novel opportunities to tailor the composition
of plant-derived lipids so that they are optimized with respect to food
functionality and human dietary needs. In order to exploit this new
capability, it is essential for food scientists and nutritionists to
define the lipid compositions that would be most desirable for various
purposes.

Document Number: 583

A variety of methods are used to analyze tomato glycoalkaloids. Because
no single method has gained wide acceptance,the extraction and analysis
of tomatine by HPLC with pulsed amperometric detection (PAD) was
previously optimized for standard and transgenic tomatoes and processed
tomato products. In the course of these studies it was discovered that
commercial tomatine contained a second glycoalkaloid, which was named
dehydrotomatine. This study demonstrates that the HPLC-PAD assay can be
used to measure both the dehydrotomatine (DT) and a-tomatine (TMT)
content of parts of the tomato plant and of low- and high-tomatine red
and green tomatoes. Both the absolute concentration of dehydrotomatine
and the percent dehydrotomatine in the mixture of the two extracted
glycoalkaloids, defined as [DT/(DT + TMT) x 100], varied widely. The DT
content of red tomatoes ranged from 0.05 to 0.42 mg/kg of fresh weight.
The corresponding range for green tomatoes was from 1.7 to 45 mg/kg. The
percent DT for the tomato plant parts was about 7 for fresh and
senescent leaves and calyxes, 10 for green fruit, 14 for small stems and
flowers, and 23 for roots and large stems. The corresponding values for
15 different tomato varieties ranged from similar to 3 to 10%. This
study demonstrates the advantages of the highly sensitive HPLC-PAD
method for DT and TMT. The possible significance of these findings for
plant and food sciences is discussed.

Document Number: 584

A new method for the selection of transgenic plants has been developed.
It is based upon selection of transgenic plant cells expressing the xylA
gene from Streptomyces rubiginosus, which encodes xylose isomerase, on
medium containing xylose. The xylose isomerase selection system was
tested in potato and the transformation frequency was found to be
approximately ten fold higher than with kanamycin selection. The level
of enzyme activity in the transgenic plants selected on xylose was 5- to
25-fold higher than the enzyme activity in control plants. Potato
transformants were stable over two generations in Southern blotting
analysis. This novel selection system is more efficient than the
traditionally used kanamycin-based selection systems. In addition, the
xylose isomerase system is independent of antibiotic or herbicide
resistance genes, but depends on an enzyme that is generally recognized
as safe for use in the starch industry and which is already being widely
utilized in specific food processes.

Document Number: 585

A method has been developed to distinguish between traditional soy beans
and transgenic Roundup Ready soy beans, i.e. the glyphosate ('Roundup')
resistant soy bean variety developed by Monsanto Company. Glyphosate
resistance results from the incorporation of an Agrobacterium-derived
5-enol-pyruvyl-shikimate-3-phosphatesynthase (EPSPS) gene. The detection
method developed is based on a nested Polymerase Chain Reaction (PCR)
procedure. Ten femtograms of soy bean DNA can be detected, while,
starting from whole soy beans, Roundup Ready DNA can be detected at a
level of I Roundup Ready soy bean in 5000 non-GM soy beans (0.02%
Roundup Ready soy bean). The method has been applied to samples of soy
bean, soy-meal pellets and soy bean flour, as well as a number of
processed complex products such as infant formula based on soy, tofu,
tempeh, soy-based desserts, bakery! products and complex meat and
meat-replacing products. The results obtained are discussed,with respect
to practical application of the detection method developed.

Document Number: 586

Methionine (Met) is the primary limiting essential amino acid in grain
legumes. The imbalance in amino acid composition restricts their
biological value (BV) to 55 to 75% of that of animal protein. So far
improvement of the BV could not be achieved by conventional breeding.
Therefore, genetic engineering was employed by several laboratories to
resolve the problem. Three strategies have been followed. A) Engineering
for increased free Met levels; B) engineering of endogenous storage
proteins with increased numbers of Met residues; C) transfer of foreign
genes encoding Met-rich proteins, e. g. the Brazil nut 2S albumin (BNA)
and its homologue from sunflower, into grain legumes. The latter
strategy turned out to be most promising. In all cases the gene was put
under the control of a developmentally regulated seed specific promoter
and transferred into grain legumes using the bacterial Agrobacterium
tumefaciens-system. Integration into and copy numbers in the plant
genome as well as Mendelian inheritance and gene dosage effects were
verified. After correct precursor processing the mature 2S albumin was
intracellularly deposited in protein bodies which are part of the
vacuolar compartment. The foreign protein amounted to 5 to 10% of the
total seed protein in the best transgenic lines of narbon bean (Vicia
narbonensis L., used in the authors' laboratories), lupins (Lupinus
angustifolius L., used in CSIRO, Australia), and soybean (Glycine max
(L.) Merr., used by Pioneer Hi-Bred, Inc., USA). In the narbon bean the
increase of Met was directly related to the amount of 2S albumin in the
transgenic seeds, but in soybean it remained below the theoretically
expected value. Nevertheless, trangenic soybean reached 100%, whereas
narbon bean and lupins reached approximately 80% of the FAO-standard for
nutritionally balanced food proteins. These results document that the
Met problem of grain legumes can be resolved by genetic engineering.

Document Number: 587

Taste-modifying proteins are a natural alternative to artificial
sweeteners and flavour enhancers and have been in use in some cultures
for centuries. Use has been limited by the stability and availability of
these proteins, but recently advances in biotechnology have been made
that will increase their availability. These include production in
transgenic organisms and protein engineering to increase stability.
Taste-modifying proteins will be available for much wider use in the
food industry, and will reduce dependence on synthetic alternatives. (C)
1998 . 

Document Number: 588

According to the << Navel Foods >> directive from the EU (February
1997), the products based on - or derived from - genetically-modified
organisms (GMO) should be labeled unless there is << substantial
equivalence >> between the GMO-derived product and the equivalent
product from non-GMO material. Currently, the most useful method to
identify, GMO's is based on the PCR technics, Methodology for transgene
detection in raw and industrially-processed materials is presented. The
limits of methodology specially for food products, the problem of
sampling, the definition of the tracing limits and the detection of
unknown transgenes are discussed.

Document Number: 589

The halobacteria are the only organisms that are tolerant of salinity at
the molecular level. All other bacteria, all fungi, all plants, and all
animals avoid the need for salt tolerance for most of their
macromolecules by maintaining defined and conserved conditions in the
cytoplasm. These conditions favour potassium over sodium, the limitation
of total inorganic ion activity, and the supplementation of this where
necessary with organic solutes which are metabolically neutral osmolytes
that may also be osmoprotectant. The salt tolerance of an organism
depends upon the range of external salinity over which it is able to
sustain these conditions in the cytoplasm. There is substantial and
increasing knowledge of the molecular biology and molecular genetics of
the processes of ion and organic solute transport, solute synthesis, and
compartmentation that underpin cell-based tolerance. Much of recent
research focuses on the identification of genes and gene products that
affect cell-based tolerance, commonly derived from single-cell models.
There is commonly the implicit or explicit assumption that incorporation
of these genes will benefit the salt tolerance of food crop species.
While this essential experimental approach is giving enormous insight
there should not be rash or premature expectations. The unique and
overriding consideration for the salinity tolerance of terrestrial
plants is the net flux of water due to transpiration and so resides at a
higher revel of organization. Processes that are advantageous to a
single cell in an aqueous medium may be lethal to a cell in a leaf in
the air. The likely impact of single structural-gene changes in ion and
solute transport upon co-ordinated plant response is probably
over-estimated, and recent Views consider regulatory processes and
multiple gene transfers. While the technical ability for plant
transformation increases daily, the practicality of using transgenic
plants in complex breeding programmes seems rarely to be given enough
thought. If intervention at the molecular level is to lead to
salt-tolerant crop plants then it will be essential to View this in the
contexts of whole plants and of plant breeding. Recent indications that
a relatively small number of quantitative trait loci (QTL) may govern
complex physiological characters offer the most hope for the future.

Document Number: 590

Due to substantial progress in breeding and cultivation practice
rapeseed has become the world's third most important source of vegetable
oil. Modification of the fatty acid composition to make rapeseed oil
more competitive in various segments of the food and industrial oil
markets has been an important objective of plant breeding and molecular
genetics in recent years. While making up the primary demand by food and
animal feed industry furnished by "double-low" quality rapeseed.
so-called "canola", interest increased to produce "Biodiesel" feedstocks
or special materials being directed to several industrial niche markets,
because of their higher value than commodity oils. Rapeseed oil is
unique in having a large spectrum of usability and good properties for
non-food applications, such as relatively homogeneous composition, high
degree of refinement, freedom from contaminants, and also
biodegradability, giving it advantages over petrochemicals.
Consequently, one of the most important objectives of rapeseed breeding
is the genetic modification of the seed oil by maximizing the proportion
of specific fatty acids, like laurate, erucate or functionalized acids,
in order to obtain tailor-made raw materials suited for industrial
purposes.

Document Number: 591

Using a variety of in vitro techniques, it is now possible to isolate a
selected gene sequence from any source and introduce it into any major
crop plant. Millions of hectares of such genetically modified (GM) or
transgenic plants are already being grown commercially, mostly in North
America. To date, the most widely grown GM crops (soybean and maize) are
those with modified agronomic traits (herbicide or insect tolerance);
the products from these commodity crops are now included in a wide range
of processed foods. This review describes the methods used to generate
these Gh I crops and then discusses the range of modified food products
that can be generated using this new technology. Such products include
those with altered protein, starch or oil quality, as well as examples
of improved micronutrient or vitamin content. Much of this work,
particularly that aiming to develop food with specific health benefits,
is still at the experimental stage, but there is no doubt that many GM
foodstuffs, with an increasing variety of qualitative changes, will
reach the market in the coming years. The rate at which such products
are developed commercially depends to a large extent on the public
reaction to a technology still poorly understood by most consumers.

Document Number: 592

The major uses of soybean oils world-wide are for food products such as
frying oils, shortenings and margarine. Refined soybean oil is usually
chemically hydrogenated to increase storage life, stability during
frying, and to increase its melting point for use in solid fat
applications. Hydrogenation results in the formation of monounsaturated
trans fatty acids, the consumption of which has been associated with an
increased risk of coronary heart disease in humans. It is nutritionally
desirable, therefore, to produce trans-free soybean oils rich in
monounsaturated fatty acids for cooking and trans-free oils rich in
stearic and oleic acids for shortenings and margarine. Cloned genes may
be introduced into soybeans to create transgenic lines with improved oil
traits. The design of transgene constructs has been assisted by the use
of soybean somatic embryos in suspension culture as a model system for
soybean seed transformation. This system has allowed the selection of
the right genes and promoters to achieve the desired phenotypes in
transgenic soybeans. By manipulating the expression of fatty acid
desaturase genes we have produced lines with 85% oleic acid in their
seed oil and lines with up to 30% stearic acid. Commercialization of
high-oleic acid transgenic soybeans has demonstrated that it is possible
to drastically alter the fatty acid composition of a soybean seed
without affecting the yield or environmental sensitivity of the soybean
plant. We have demonstrated that high-oleic soybean oil is also useful
for non-food applications such as biodegradable lubricants. We have now
cloned a number of fatty acid desaturase-related genes from species
which produce unusual and industrially useful fatty acids. By expressing
these genes it will be possible to produce new fatty acids in soybean
seeds, which could potentially replace petrochemicals as raw material
for many industrial processes.

Document Number: 593

The microbial bar and pat genes confer tolerance to the non-selective
herbicide phosphinothricin (PPT; sold as Basta or Finale). This
tolerance in plants could provide an environmental gain compared to
current-day herbicide cocktails, but the safety of such a transgene
approach is questioned by many. The biosafety of the presence of these
herbicide tolerance genes in plants is evaluated in a
'transgene-centered approach'. Potentially, the introduction of
transgenic PPT-tolerant crops could result in acquired PPT tolerance in
weedy relatives of these crops. Assuming responsible use of this trait
in agronomy, the ecological consequences with respect to weediness or
spread of the transgenic PPT tolerance are concluded to be negligible.
The key issue for the toxicological evaluation is whether or not the
plant has actually been sprayed with PPT. Consumption of the gene and/or
gene product from unsprayed transgenic plant material will not have
adverse effects. In case of PPT-sprayed material, PPT or its derivatives
could be present in food and feed and crop-specific metabolites might be
formed. To date, the toxicological impact of such a putative exposure is
not sufficiently clear, and further premarket testing is recommended.

Document Number: 594

The genetically modified corn 'MaisGard(TM)' (Monsanto, 'YieldGard(TM)'
in the U.S.A.) is resistant to the European corn borer due to the
insertion and expression of the cryIA(b) gene from Bacillus
thuringiensis ssp. kurstaki. This gene controlled by a 35S promoter from
the cauliflower mosaic virus is flanked at its 5'-site by the
corn-specific intron 1 of heat shock protein 70 (hsp 70). MaisGard(TM)
corn is already widely cultivated in the U.S.A. and may find its way to
Europe as a component of a variety of food- and feedstuffs. This work
describes the development and evaluation of a sensitive and specific
detection method for MaisGard(TM) corn using a nested polymerase chain
reaction (PCR) system. The system consists of an outer 401 bp fragment
spanning the region of the 35S-promoter to the hsp70 intron 1, and an
inner 149 bp fragment. The method was developed using a gradient cycler
to empirically determine the range and limits of annealing temperatures
that can be used. The sensitivity of the method is 50 pg DNA when using
pure transgenic DNA, and 500 ng DNA when 0.01% of the analysed DNA is
transgenic. No other tested transgenic corn, including Bt176 and Bt11
from Novartis and 'LibertyLink(TM)' (T25) from AgrEvo or conventional
corn could be detected with the MaisGard(TM)-system, indicating the high
specificity of the method for MaisGard(TM).

Document Number: 595

Two potato varieties, Rustica and Desiree, were tested for a genetic
modification consisting of a granule bound starch synthase (gbss) gene
in antisense orientation combined with the endogenous B33 promoter and
parts of the pBIN 19 plasmid vector including the neomycin
phosphotransferase II (nptII) gene as a marker. Various polymerase chain
reaction (PCR) primers were constructed for vector sequences, target
inserts and the marker gene. At first no products with the predicted
target-insert size were obtained. However, a long-template PCR combined
with a nested PCR led to the expected amplification product in the
Desiree variety. Results were confirmed by a restriction endonuclease
digest, Southern blotting of fragments and by biochemical tests. The
potato variety Rustica showed no target insert sequences on molecular
analysis, nor did biochemical methods indicate modifications of the
phenotype (i.e. reduced amounts of gbss protein and amylose). Only
vector sequences and the marker gene were detected by the PCR method.

Document Number: 596

Assessing the potential allergenicity of transferred proteins remains
one of the most difficult aspects of the overall safety assessment for
transgenic foods. Allergenicity assessment must consider several
factors, including the source of the transferred protein, expression
levels, the physical and chemical properties of the protein, and
similarity to known allergens. Although no single factor can be
considered definitive, consideration of all these factors together may
provide some indication of potential allergenicity.

Document Number: 597

Development of new industrial plants is one of the main purposes of
modern biotechnology. Whereas transgenic oilseed crops have entered the
state of agroindustrial exploitation, modification of plant carbohydrate
metabolism is still in its infancy. Despite of its role as one of the
worlds most important crops and a high potential of sugar and sugar
derivatives as food ingredients and precursors of economically valuable
substances sugarbeet has become amenable to metabolic engineering only
slowly. The most serious problems have been imposed by inefficient and
poorly reproducible regeneration protocols and an almost complete lack
of cloned genes known to be expressed specifically in the storage root.
The current state of transformation technology suggests that many
metabolic alterations of sugar beet root metabolism can be introduced
into the plant, if a cloned gene is available providing the enzyme
function needed to catalyze the required modification step. Examples of
this category are elimination of undesired byproducts like raffinose,
invert sugar and dextran, one step synthesis of simple sugar
derivatives, e.g. ketosugars or fructans, and also production of ethanol
and possibly other products of microbial fermentations by post harvest
induction of the necessary enzymes. Among the more distant breeding
objectives are the synthesis of better pectins and lignin free cellulose
in sufficient amounts and quality to allow technical applications. This
will be feasible only when most or all the genes involved in synthesis
of these biopolymers will be known. Transformation and regeneration of
sugarbeet has now been improved significantly. Moreover, effective
molecular methods are available for isolation of developmentally
regulated genes. Molecular breeding of a first generation of
metabolically engineered sugar-beet cultivars should thus take no longer
and not be more difficult than in the case of other transgenic crops
like oilseed rape, that are now used commercially.

Document Number: 598

Russet Burbank potato was transformed with plant expression vectors
containing the potato leafroll luteovirus (PLRV) coat protein (CP) gene.
Transgenic potato lines contained a gene expression cassette with two
copies of a PLRV CP gene in which the nucleotide sequence was modified
to improve expression of the gene. In addition, the two copies of the
PLRV CP gene were each driven by a different promoter. Field test
screening for PLRV resistance identified 15 lines which showed moderate
resistance to PLRV infection and virus titer build-up and a longer
incubation period for systemic infection. By conducting field resistance
assays during a period when the vector of PLRV was not present, it was
possible to test whether the observed resistance was sufficient to
restrict aphid transmission of PLRV in a field test. Two years of field
testing demonstrated that PLRV-spread from an infected plant to adjacent
healthy plants of the same line was severely restricted in nearly all
the transgenic lines in the field. These lines have useful resistance to
PLRV and could aid in managing PLRV disease in Russet Burbank potato.

Document Number: 599

Recombinant DNA technology provides a powerful tool to create new
products in many different fields. Agriculture and the agro-food
industry are two of them. Genetic engineering aims at improving the
yield, the nutritional quality or the technological value of food crops.
The composition of foods may also be modified in order to decrease the
content of toxic substances sometimes present in certain food crops. The
expression of recombinant proteins in foods must be carefully assessed
as proteins may induce allergic reactions in humans. Currently, the
potential allergenicity of a protein can be reasonably assessed only
when the protein is known to trigger an immune response in sensitive
subjects. By contrast, the potential allergenicity of a protein of
unknown allergenicity cannot be easily predicted as no immunoserum of
allergic subjects is available. That is why an allergenicity assessment
model for any genetically engineered food should be designed. Even
though the gene product is completely characterised in the transgenic
product, this does not necessarily provide information on its potential
allergenicity. Animal models, in vitro tests and protein structure
should be taken into account. Besides, allergenicity is a biosafety
issue, for it deals with human health. The issue of food allergies in
food safety should not be neglected, as an allergic reaction can be life
threatening.

Document Number: 600

To assess the knowledge and attitude of Canadian dietitians towards
biotechnology we moiled a 50 item, validated questionnaire to 1,000
English-speaking members of Dietitians of Canada in the Fall of 1996.
Knowledge questions tested regulatory polities, genetics and transgenic
foods. Using a 5-point Likert stale, attitude questions addressed four
areas: role of the dietitian, science and agriculture, safety and
labelling of transgenic: foods and nutritional quality. Demographic
information was also obtained. The response rate was 45.6 %. There were
no significant gender, location or area of practice differences.
Overall, knowledge and attitude were significantly correlated (r = 0.16,
P < 0.0001). Knowledge correlated with attitudes concerning safety and
nutrition. Attitude hut not knowledge was significantly lower in British
Columbia (n=56) compared to Atlantic (n=76), Central (n=190) or prairie
(n=124) regions. Dietitians who had never received any information on
biotechnology stored lower than the rest of respondents on knowledge and
attitude toward the role of the dietitian and safety. The good response
rate to this survey, the favourable attitude to the role of the
dietitian and an expressed desire to have further access to information
on biotechnology, suggest dietitians believe biotechnology to be an
important issue; however, access to information about new products is
needed.

Document Number: 601

The quality of tomato puree prepared from transgenic fruits
(Lycopersicon esculentum) which had reduced amounts of polygalacturonase
(PG) activity, were evaluated. The application of genetic modification
yields products which have lower Bostwick consistency values (from 35 to
55%), reduced serum separation (c. 40%) and better sensory appearance
and acceptability in comparison with their conventional counterparts. A
new way of producing high quality tomato products using genetically
modified fruits together with mild heat treatments is described.

Document Number: 602

The first generation of genetically modified crops are making their way
towards large-scale release and the market. Small scale trials have been
made world-wide and assessments of field safety attempted. This review
summarises discussions of non-food issues arising from common types of
construct marker genes mostly antibiotic resistance and herbicide
tolerance and same genes expressing resistance to insects. Some
herbicide tolerant potato and beet present risks for future crops.
Continuing observation of early trials is recommended, particularly for
effects on pollinators. Virus inserts need further study but only using
genes already common to the area helps minimise risk. Trees and grasses
and same anti-pest genes may present future problems. There is a
detailed locus on potato, sugar beet and oil seed rape as agricultural
weeds and on their gene flow to weeds and feral forms and further study
is recommended Weediness, invasiveness and risks to the nonagricultural
environment and risks from virus coat protein genes inserted in crops
are briefly reviewed.

Document Number: 603

Biotechnology includes the application of a wide variety of biological,
biochemical, bioengineering, genetic, microbiological and control
techniques. The baking of yeast-leavened and sourdough breads represents
one of the oldest biotechnical processes, together with the brewing of
beer, sake and wine, and the production of yoghurt and cheese, etc. A
modern baking process may take advantage of biotechnology in its widest
sense, from the improvement of cereal grains and starter cultures by
recombinant DNA technology, through the use of enzymes as processing
aids, to application of the most advanced batch and continuous
fermentation technologies.

Document Number: 604

Wheat is the number one food crop in the world based on area under
cultivation and total production. The popularity of wheat is based on
diverse uses, resistance to many pathogens and pests, low cost of
production, rapid growth, genetic flexibility and adaptability to
different climates. The unique breadmaking quality is related to the
type and quantity of gluten proteins, especially the
high-molecular-weight glutenin subunits that are synthesized and stored
in the seed endosperm. Recent advances in genetic transformation of
wheat, including the integration and expression of high-molecular-weight
glutenin subunit genes, now make it possible to engineer the gluten
proteins in order to improve breadmaking qualities.

Document Number: 605

This paper summarises the key issues raised at a recent international
conference entitled, 'Integrated Crop Protection: Towards
Sustainability?' The British Crop Protection Council in association with
the Sustainable Farming Systems initiative - a joint enterprise between
the Scottish Agricultural College and the University of Edinburgh -
organised during 1995 an international conference to address
state-of-the-art studies in the theory and practice of weed, pest and
disease management by integrated approaches. The conference also
explored the contribution which sustainable fanning systems research can
make to mainstream agriculture. The major recurring themes were: (i) are
current crop protection practices sustainable?; (ii) the concept of
sustainability; (iii) the biological roots of sustainability; (iv)
anticipated developments in crop protection and sustainability; (v) food
production and sustainability; (vi) experimentation on sustainable
farming systems; and, finally, (vii) are integrated crop protection
practices moving towards an increased sustainability? The key issues
raised in theme (i) were: crop protection is of critical importance to
all food production systems; and the way we protect our crops today has
significant implications for future generations; theme (ii): the concept
of degrees of sustainability, i.e. sustainability not as an absolute
concept but with the potential for systems to be more or less
sustainable; theme (iii): the essential nature of developing holistic
knowledge of systems, and not simply by discipline only; theme (iv): the
development of decision support technology, transgenic plants and
semiochemicals as tools in future crop protection strategies; theme (v):
the need for continued government funding to meet the twin, and often
conflicting, demands of food production and sustainability; and themes
(vi) and (vii): more emphasis must be given to systems experimentation
and to technology transfer, respectively. (C) 1997 Elsevier Science B.V.

Document Number: 606

Fructans (polyfructosylsucrose) are synthesized by a number of plants
and micro-organisms. Plant fructans are localized in the vacuole and
have a low degree of polymerization (DP), whereas the fructans
synthesized by microorganisms are usually much bigger. There is an
increasing interest in fructans for both food and non-food applications.
In order to accumulate fructans of high DP in the plant vacuole, the
levansucrase protein of Bacillus subtilis was fused to the vacuolar
targeting sequence of sporamin and expressed in plants. Transgenic
tobacco plants in which this fusion gene is expressed accumulate
fructans to levels up to 21% of the d. wt. They showed a reduced
translocation of carbohydrates, bleaching of the leaves, stunted growth
and increased levels of hexoses and starch. The levansucrase protein was
not translocated to the plant vacuole, but retained in the endomembrane
system, even though the same targeting signal was able to translocate
the E. coli GUS protein to the plant vacuole.

Document Number: 607

Rice (Oryza sativa L.), the major food staple for more than two billion
people, contains neither beta-carotene (provitamin A) nor C-40
carotenoid precursors thereof in its endosperm. To improve the
nutritional value of rice, genetic engineering was chosen as a means to
introduce the ability to make beta-carotene into rice endosperm tissue.
Investigation of the biochemical properties of immature rice endosperm
using [C-14]-labelled substrates revealed the presence of geranyl
geranyl diphosphate, the C-20 general isoprenoid precursor necessary for
C-40 carotenoid biosynthesis. Phytoene synthase, which condenses two
molecules of geranyl geranyl diphosphate, is the first of four specific
enzymes necessary for beta-carotene biosynthesis in plants. Therefore,
the Japonica rice model variety Taipei 309 was transformed by
microprojectile bombardment with a cDNA coding for phytoene synthase
from daffodil (Narcissus pseudonarcissus) under the control of either a
constitutive or an endosperm-specific promoter. In transgenic rice
plants, the daffodil enzyme is active, as measured by the in vivo
accumulation of phytoene in rice endosperm. Thus, it is demonstrated for
the first time that it is in principle possible to engineer a critical
step in provitamin A biosynthesis in a non-photosynthetic,
carotenoid-lacking plant tissue. These results have important
implications for longterm prospects of overcoming worldwide vitamin A
deficiency.

Document Number: 608

Wheat gluten proteins are largely responsible for the viscoelastic
properties that allow doughs to be processed into bread and various
other food products including cakes, biscuits (cookies), pasta and
noodles. Detailed biochemical and biophysical studies are revealing
details of the molecular structures and interactions of the individual
gluten proteins, and their roles in determining the functional
properties of gluten. In particular, one group of gluten proteins, the
high molecular weight (HMW) subunits of glutenin, have been studied in
detail because of their role in determining the strength (elasticity) of
doughs. The development of robust transformation systems for bread wheat
is now allowing the role of the HMW subunits to be explored
experimentally, by manipulating their amount and composition in
transgenic plants. Such studies should lead to improvement of the
processing properties of wheat for traditional end uses and the
development of novel end uses in food processing or as raw material for
other industries.

Document Number: 609

The first food products derived from transgenic plants that are
resistant to diseases, insects or viruses are now reaching the market
and there is growing public concern about problems of allergenicity and
toxicological changes in such transgenic food plants. We review the
modifications being carried out or envisaged in molecular resistance
breeding and specifically consider the allergenic and toxicological
potential of the gene products used. Several protein Families that
contribute to the defence mechanisms of food plants have members which
are allergens or putative allergens and some of these proteins are used
in molecular approaches to increase resistance. These include
alpha-amylase and trypsin inhibitors. lectins and pathogenesis-related
proteins. An assessment procedure to avoid the transfer of such
allergens is described. The source of the transgene is of great
importance for the application of immunological assays. In addition to
putative changes in the allergenic potential, the toxicological
implications of classical and molecular resistance breeding are
discussed. Several 'self defence' substances made by plants are highly
toxic for mammals, including humans. Examples of molecular approaches
that could be of toxicological concern are given. The source of the
transgene is of no relevance in assessing the toxicological aspects of
foods from transgenic plants. Food safety can also be severely
influenced by invading pathogens and their metabolic products. This may
result in a trade-off situation between 'nature's pesticides' produced
by transgenic plants or varieties from traditional breeding programmes,
synthetic pesticides and mycotoxins or other poisonous products of
pests.

Document Number: 610

In 1994, the first genetically modified plant was approved for
cultivation and commercialization as a foodstuff in the USA. Now, at
least fifteen genetically modified plants have been approved
additionally. In Canada and other countries further applications for
transgenic plants are pending. Altough the regulatory frameworks
concerning the commercialization of Novel Food differ within EC, foods
derived from genetically modified plant varieties (i.e. rapeseed,
radicchio, soybeans, tobacco) have been introduced into commerce in
Europe. At present an appropriate labelling of foods developed by means
of new DNA techniques is still being discussed. Consequently, approaches
for detection of DNA in foods derived from genetically modified
organisms are necessary to control the correct labelling of foods and
for the safety of producers and consumers. For the detection of
transferred DNA the polymerase chain reaction (PCR) or hybridization
techniques can be used. A prerequisite for detection is a minimum number
of copies and the information about the sequence of the introduced
genetic material. DNA sequences which are responsible for the new traits
(structural genes, i.e. for herbicide or virus resistance) and which do
not occur naturally in the host plant, are necessary for the specific
detection. But the detection can also be performed with genes which
funktion as control elements or marker genes (promotor, terminator,
nptll-gene). This paper describes a screening method for the detection
of six different transgenic plants by the polymerase chain reaction
(PCR). The presence of the transferred control elements were determined
with primer pairs specific for the CaMV 35S promotor, NOS terminator
(Nopalin synthethase) and the nptll gene (Neomycinphosphotransferase).

Document Number: 611

Fungicides continue to be essential for the effective control of plant
diseases. New classes of fungicides with novel modes of action are being
developed in the 1990s. These include the strobilurins, phenylpyrroles,
anilinopyrimidines, phenoxyquinolines, and compounds that trigger
defense mechanisms in the plant. For the foreseeable future, new
toxophores will be identified through a process of random screening,
with natural products representing a rich source of fungicide leads.
Progress is being made in the development of high-throughput screens
comprised of target enzyme sites or cell-based assays; these techniques
will improve the probability of discovery. Following the identification
of suitable leads, biorational design is used to optimize specific
properties. In vivo glasshouse screens and field trials are expected to
remain the dominant methods for characterizing new compounds. Low
toxicity to humans and wildlife, low environmental impact, low residues
in food, and compatibility with integrated pest management (IPM)
programs are increasingly important considerations in the selection of
fungicides for development.

Document Number: 612

Scientific views on the importance and potential risks of plant genetic
engineering are summarized and the ecological and economic consequences
of the large scale release of engineered crops is discussed. The
difference between scientific risk assessment and the public's
perception of risk due to modification of the former by outrage factors
is explained. Polls, media, and literature sources show that the public
supports plant genetic engineering activity, but has some concerns.
These concerns fall within two categories: (1) it is risky, (2) it is
morally wrong. Polls show a lack of knowledge about GE by the public and
that outrage factors play a major role; their effects on perception are
described in detail. Ethical considerations also exercise a powerful
influence over GE's acceptance, but the complementary roles of Religion
and Science are emphasized. Different attitudes to nature are described
and it is concluded that if the world is to be fed and life support
systems sustained, active management of nature is required.
Lastly, the need for scientists to communicate with the public and the
basis of effective presentation is discussed.

Document Number: 613

Melon (Cucumis melo L.) is an important fruit crop cultivated widely in
every region of the world. Our laboratory is targeting this species for
production of novel proteins important to food industry. Prior to
expression of protein of interest in transgenic melon an efficient
genetic transformation system has to be developed. In this context we
are testing a wide variety of promoters fused to reporter gene for
beta-glucuronidase (GUS) for expression specifically in melon fruits. In
this study in melon, salicylic acid-inducible promoter region of
pathogenesis-related protein gene (PR1a) of tobacco fused to
beta-glucuronidase (GUS) gene was introduced into melon via
Agrobacterium-mediated gene transfer using a binary vector system. Gene
transfer was effective when Agrobacterium virulence factors like
acetosyringone (100 mu M) and low pH (5.2) were provided during the
co-culture step. Transformed shoots were recovered from
benzyladenine-induced cut cotyledons using kanamycin gene as a selective
marker. Regeneration of shoots from cotyledons was stimulated by
providing 10 mM proline in the shoot organogenesis medium. Southern and
Northern blot analysis of transformants confirmed the presence of
beta-glucuronidase gene in two selected clones J-3 and PR-G. The
transformants also showed high beta-glucuronidase activity after
salicylic acid treatment. Thiamine, a previously known inducer of
pathogenesis-related protein, stimulated beta-glucuronidase in J-3 but
not PR-G melon transformants tested in this study. These studies showed
that tobacco PR1a promoter region can be expressed in melon and it was
stimulated by salicylic acid. This indicates the potential to use the
promoter region of tobacco PR1a for genetic improvement of melon for
specific food processing-related characteristics or for expression of
novel food-related proteins. The promoter region could be used to drive
specific target genes under stress or salicylic acid induced conditions.

Document Number: 614

Allergenic proteins with a molecular mass of about 14 to 16kDa were
isolated from a rice salt-soluble fraction based on the reactivity with
IgE antibodies from patients allergic to rice. cDNA clones encoding
these allergenic proteins were isolated from a cDNA library of maturing
rice seeds, and the deduced amino acid sequences showed considerable
similarity to wheat and barley alpha-amylase/trypsin inhibitors, which
have recently been identified as major allergens associated with baker's
asthma. An antisense RNA strategy was applied to repress the allergen
gene expression in maturing rice seeds. Immunoblotting and ELISA
analyses of the seeds using a monoclonal antibody to a 16-kDa allergen
showed that allergen content of seeds from several transgenic rice
plants was markedly lower than that of the seeds from parental wild type
rice.

Document Number: 615

Lipid metabolism in plants provides uncommon opportunities for genetic
engineering to produce plant oils suited to a variety of end-uses. These
opportunities include improvement of food and nutritional value,
creating specialty lipids and feedstocks for high-value products and
designing custom-made materials for industry. Genetic engineering
intervention for production of novel transgenic plants which elaborate
the desired product has graduate from academic exercise to commercial
possibilities. It is now realized that transgenic crops can serve as
biological factories for upscaling production of premium lipids via
molecular farming. This review is an attempt at analyzing the status in
this field.

Document Number: 616

Transformation and regeneration is routine for many crop plants. A
genetically engineered tomato with a longer shelf life at full ripeness
was introduced in the United States in 1994, and other
soon-to-be-released products, both foods and fibers, incorporate genes
for resistance to pests, diseases, and environmentally benign
herbicides. Other possibilities are altered plant fats and oils,
methionine- and lysine-enhanced grain and legume proteins, plant foods
that can deliver immunizing antigens, and other ways of controlling
fruit ripening. Food safety concerns include the inadvertent production
of toxicants and allergens. Foreign DNA can be introduced into plants by
bacterial vectors, direct uptake by protoplasts, and mechanical
introduction on metal particles or other materials. Limitations include
little or no control of copy number or site of integration of the
introduced DNA, dependence on selectable markers far recovery of traits,
and inadequate knowledge of how to control key metabolic steps to
maximize desirable traits. Directed genetic change still requires
conventional crop breeding to deliver benefits to farmers and consumers.

Document Number: 617

Two studies, one with four cultivars grown at four locations during 1989
to 1993, and the second with a transgenic and its non-transformed parent
cultivar grown at three locations for three years, were undertaken to
explore the variation in total flavonoid content of flaxseed (Linum
usitatissimum). Results of the analyses of variance for the first study
showed significant cultivar as well as environmental effect on flavonoid
content. Cultivar NorLin displayed the highest flavonoid content across
the sites with a mean of 71 mg/100 g, and Omega, a yellow-seeded
flaxseed cultivar, the lowest with a mean of 35 mg/100 g. There was no
significant difference between a transgenic flaxseed and its
non-transformed parent for total flavonoid content except for one
location in 1991. Flavonoid content was inversely related to the protein
content and there was a weak positive correlation between flavonoid and
oil levels.

Document Number: 618

In recent years, a number of agricultural crops have been developed with
recombinant DNA technology. Because the transferred genes code for
proteins that are ordinarily not present in these particular foods,
there is concern about the potential allergenicity of these new crop
varieties. Foods contain many proteins; however, only a small fraction
are allergens. Although the structural properties of proteins that cause
allergic reactions have not been characterized completely, known food
allergens in general have molecular weights between 10 and 70 kDa,
stimulate the immune response (induce the production of
allergen-specific IgE), and are stable molecules that are resistant to
processing, cooking, and digestion. Although any type of food is
potentially allergenic, the majority of food allergies are caused by a
small group of foods (cows' milk, nuts, legumes, eggs, seafood).
Cross-reactivities occur within a given food group and between foods and
seemingly unrelated proteins. Even though most transgenic foods are
considered safe, biotechnological manipulation can affect crop
allergenicity. The safety evaluation of transgenic foods is relatively
easy when the allergenicity of the gene sources are known. The
recombinant food can be assayed using traditional in vitro inhibition
assays. Recently, reduced allergen content of biotechnologically altered
rice was shown. In contrast, increased allergenicity was demonstrated in
transgenic soybeans after a methionine- and cystine-rich protein from
Brazil nuts, identified as a major Brazil nut allergen, was expressed in
soybean to increase its content of sulfur-rich amino acids. The most
difficult issue regarding transgenic food allergenicity is the effect of
transfer of proteins of unknown allergenicity. The challenge is to
determine whether these proteins are allergenic as there is no generally
accepted, established, definitive procedure to define or predict a
protein's allergenicity. Comparing the structures of the transferred
protein with known allergens and allergen epitopes could be one
approach. Additionally, Th-2 cell stimulation, the production of IL-4,
and IgE antibody induction in animal models may help to evaluate the
potential allergenicity of a protein.
In conclusion, there is no evidence that recombinant proteins in newly
developed foods are more allergenic than traditional proteins. The
evidence suggests that the vast majority of these proteins will be
completely safe for the consumer. The concern is that if a few
transgenic foods cause serious allergic reactions, this could undermine
the public's confidence in such products. It is essential that proper
guidelines are established and tests are developed to assure that this
will not occur.

Document Number: 619

The major use of the ten billion pounds or so of soybean oil produced in
the US is for food products such as cooking oils, shortenings and
margarines. Refined, bleached and deodorized (RED) soybean oil used for
cooking is usually hydrogenated to increase storage life and stability
during frying. RED soybean oil is also extensively hydrogenated to
increase melting point for functionality in shortenings and margarines.
Hydrogenation results in oils rich in trans fatty acids, the consumption
of which may be associated with coronary heart disease. RED oils used
for salad oils are not hydrogenated but are rich in palmitic acid, the
consumption of which has also been associated with coronary heart
disease. Therefore, it is nutritionally desirable to produce trans-free
soybean oils rich in monounsaturated fatty acids with reduced palmitic
acid for cooking and soybean oils with no saturated fatty acids for
salad oils. It is also desirable to produce trans-free oils rich in
stearic and oleic acids for shortenings and margarines. Cloned genes may
be introduced into soybeans to create transgenic lines with improved oil
traits. The design of transgene constructs has been assisted by the use
of soybean somatic embryos in suspension culture as a model system for
soybean seed transformation. This system has allowed the selection of
the right genes and promoters to achieve the desired phenotypes in
transgenic soybeans. Current soybeans in development include lines
producing oil with reduced palmitic acid, lines with over 80% oleic acid
and lines with up to 30% stearic acid. Commercialization of high oleic
acid transgenic soybeans has demonstrated that it is possible to
drastically alter the fatty acid composition of a soybean seed without
affecting the yield or environmental sensitivity of the soybean plant.

Document Number: 620

Novel products can be created by instant packaging oi enzymes in
transgenic seeds using plant genetic engineering technique. The
seed-formulated enzyme preparation is convenient for handling, and very
stable during storage. The transgenic seeds can be applied without any
purification, in the leed, food and raw materials processing industries.

Document Number: 621

Recombinant DNA technology and techniques for gene insertion into
animals have generated promises of animals genetically engineered to be
healthy, productive, and sources of novel and/or improved foods and
fibre, Transgenic animals have also generated concerns about the
production of monsters that will cause ecological catastrophes. Here,
current research on the insertion of new genes into livestock is
described, and an attempt is made to provide a scientific perspective on
the likelihood of either outcome.

Document Number: 622

Cassava is the most agronomically important of the cyanogenic crops,
Linamarin, the predominant cyanogenic glycoside in cassava, can
accumulate to concentrations as high as 500 mg kg(-1) fresh weight in
roots and to higher levels in leaves, Recently, the pathway of linamarin
synthesis and the cellular site of linamarin storage have been
determined, In addition, the cyanogenic enzymes, linamarase and
hydroxynitrile lyase, have been characterized and their genes cloned.
These results, as well as studies on the organ- and tissue-specific
localization of linamarase and hydroxynitrile lyase, allow us to propose
models for the regulation of cyanogenesis in cassava, There remain,
however, many unanswered questions regarding the tissue-specific
synthesis, transport, and accumulation of cyanogenic glycosides. The
resolution of these questions will facilitate the development of food
processing, biochemical and transgenic plant approaches to reducing the
cyanogen content of cassava foods.

Document Number: 623

Tomato plants (Lycopersicon esculentum) synthesize the glycoalkaloid
a-tomatine, possibly as a defense against insects and other pests. As
part of an effort to improve the safety of plant foods, the usefulness
of a new HPLC pulsed amperometric detection (PAD) method for the direct
analysis of cx-tomatine in different parts of the tomato plant; in
store-bought and field-grown, including transgenic, tomatoes; in a
variety of commercial and home-processed tomato products; and in
eggplant and tomatillos was evaluated. The method was found to be useful
for analysis of a variety of products including high-tomatine calyxes,
flowers, leaves, roots, and stems of the tomato plant (14-130 mg/100 g
of fresh weight), low-tomatine red tomatoes (0.03-0.08 mg/100 g),
intermediate-tomatine tomatoes (0.1-0.8 mg/100 g), and high-tomatine
fresh and processed green, including pickled and fried, tomatoes (0.9-55
mg/100 g). No experimental difficulties were encountered with extraction
and analysis of tomatine in complex foods such as tomato juice, ketchup,
salsa, sauce, and sun-dried tomatoes. Microwaving and frying did not
significantly affect tomatine levels of tomato foods. The tomatine
content of fresh market and transgenic delayed-ripening varieties was
not different from the range ordinarily seen in tomato. The possible
usefulness of the findings to plant science, food safety, and human
health is discussed.

Document Number: 624

The functional properties of wheat flour in various food systems (most
notably for breadmaking) are determined largely by the structures and
interactions of the prolamin storage proteins, termed gliadins and
glutenins. It is therefore important to understand the structures of the
individual proteins, their pathways and mechanisms of synthesis and
deposition, and their interactions with other flour components (for
example, starch and lipids) in order to manipulate the quality of wheat
for food and other uses, Me are using a range of biophysical and
biochemical approaches to determine the structures of individual
proteins, using fractions purified from flour or expressed in
heterologous systems. We are also isolating cDNAs for proteins involved
in protein folding and assembly in the endoplasmic reticulum (protein
disulphide isomerase, BiP and peptidyl-proly cis-trans isomerase) and
studying their interactions with gluten proteins synthesised in E. coli.
This work is complemented by immunocytochemical studies of gluten
protein deposition in the leaves and seeds of transgenic tobacco plants,
using wild-type proteins and mutants in which specific transit-targeting
sequences have been added or deleted.
Taken together these results are giving a detailed picture of the
structure, synthesis, assembly and deposition of wheat gluten proteins.

Document Number: 625

The use of transgenic plants in breeding makes it possible to utilise a
wide variety of novel genes from unrelated plants, microbes and animals.
Because of the diversity of genes that have now become available for
modifying crop plants, it is agreed internationally that there should be
a risk assessment before transgenic plants are grown outside the
laboratory or glasshouse. Various aspects are considered in a risk
assessment including any non-target effects of the transgene, changes in
plant persistence and invasiveness, and the possibility of movement of
the transgenes to wild populations by cross pollination. It is generally
argued that the need for risk assessment and regulation should be
determined by an analysis of certain products of transformation, rather
than a risk assessment being required for all plants modified by the
process of transformation. A possible consequence of considering the
product only, however, could be that some of the products of
conventional breeding may need to be assessed by the risk assessment
procedures developed for transgenic plants. There are discussions with
interest groups on the use of transgenic plants in the environment and
in food products. It is likely that some form of labelling will be
required for certain foods containing ethically-sensitive genes. There
is little doubt that transgenic plants will make a significant
contribution to agriculture in the coming decades. Developments in the
patenting of genes, release regulations, food labelling, consumer
reaction etc., will influence the rate of progress and should be
considered in the strategic planning of plant breeding programmes.

Document Number: 626

Indica-type rice provides the staple food for two billion people in
Third World countries. Several problems involved in the stable and
sustained production of high quality food cannot be solved by
traditional breeding. Methods have been established for gene transfer to
Indica rice breeding lines to study possible contributions from genetic
engineering. Experiments are in progress on the development of
transgenic resistance towards Yellow Stem Borer, resistance towards Rice
Tungro Virus, accumulation of provitamin A in the endosperm, increase of
essential amino acids in the endosperm such as lysine, cysteine and
methionine and resistance towards fungal pests such as Rice Blast and
Sheath Blight. Transgenic clones from Indica rice breeding lines have
been recovered from several of the approaches mentioned, some of which
have been regenerated to plants.

Document Number: 627

The development in molecular biology over the past decade has provided
the means for transferring DNA from one species to another. Methods used
in microorganisms, plants and animals are briefly described.
Today it is possible to create herbicide-resistant plants, crops with
improved food quality and transgenic crops producing novel compounds e.
g. pharmaceuticals, substances to all industrial sectors. In the near
future it is likely to produce plants with drought and salt tolerance
and more genetically complex traits, such as nitrogen fixation.
Attributes such as disease and pest resistance, ripening
characteristics, nutritional composition, palatability, texture and
shelf life are being manipulated in ways never before possible.
This review updates several experiments in transgenic animals to alter
rate of growth, carcass composition and new technology with the goal of
producing, large amounts of valuable human pharmaceutical drugs in a
more efficient manner.
Some of the applications and implications that this technology has for
each stage of the 'food chain' - from the agricultural production of raw
materials, through processing,storage and distribution, to the labelling
of finished products - are discussed.

Document Number: 628

The thaumatins are a class of intensely sweet proteins isolated from the
fruit of the tropical plant Thaumatococcus danielli. Thaumatin is
approved for use in many countries and has application as both a flavor
enhancer and a high-intensity sweetener. The supply of naturally
occurring thaumatin is limited, which has prompted extensive research
into its synthesis via transgenic organisms. The gene encoding thaumatin
has been introduced into various microorganisms under transcriptional
control of heterologous promoters. Yields to date have been low, but the
factors governing more efficient microbial production have been
identified. Continued research should allow microbial yields to be
improved to commercially viable levels. The unique properties of
thaumatin as a food additive could well be exploited by the food
industry. Alternatively, the thaumatin gene could be engineered directly
into selected fruit and vegetable crops to improve their flavor and
sweetness.

Document Number: 629

The presence of marker genes in genetically modified crop plants raises
a number of food safety issues. Recently, the demonstration that these
genes can be removed after successful genetic modification has renewed
the discussion on their presence. Here, the potential detrimental
effects of marker genes on food safety are discussed; consideration of
these should be included as part of the risk-benefit analysis of
genetically modified food products before their introduction to the
marketplace.

Document Number: 630

Domestic farm animals play an important role in meeting some basic needs
of humankind, especially food and clothing. The aspects of genetic
improvement programs in livestock production pertinent to animal welfare
and animal well-being are reviewed. A link is made between the
evolutionary processes of adaptation and domestication and animal
well-being. Animal behavior is a component of all these. Thus, the
genetics of behavior may provide clues to the well-being of farm animal
populations, and it will also be of relevance to public opinion issues
of animal welfare. Many expressions of behavior by domestic livestock
may be influenced by those processes that change gene as well as
genotypic frequencies such as inbreeding, drift, and artificial
selection. The environment in which the individual lives will also play
a role, along with the interaction between genotype and environment.
Selection for or against such behaviors as aggressiveness, docility,
response to stress, and certain sexual behaviors in some livestock
species has often been successful. This points to the existence of
additive genetic variation for behavior, and scope for the inclusion of
behavioral traits into selection programs, if these measures are shown
to be related to welfare. Negative relationships between behaviors
associated with well-being and traits of economic importance have been
reported in most livestock species. However, estimates of genetic
parameters, especially genetic correlations between objective measures
of well-being and production traits, are scarce. There have been no
comprehensive studies of the welfare of transgenic animals reported in
the scientific literature. Increased use of biotechnology in animal
agriculture, coupled with greater public scrutiny of livestock
industries, may precipitate decisions concerning the interface of
behavior and genetics that need to be addressed before scientists can
conduct appropriate experimental evaluations.

Document Number: 631

Calgene conducted a thorough review and analysis of the use of the
kan(r) gene and gene product, aminoglycoside 3'-phosphotransferase II
(APH(3')II), for use as a selectable marker in FLAVR SAVR(TM) tomatoes,
in BXN(TM) cotton and oil-modified rapeseed. The data generated support
the conclusions that APH(3')II is not a toxin or allergen, that the ka
nr gene is highly unlikely to move from the plant genome into
microorganisms via horizontal gene transfer, that if such transfer could
occur the impact would be minimal, and that APH(3')II in transgenic
plants will not compromise antibiotic use in humans or animals.
At FDA's Food Advisory Committee meeting on April 6-8, 1994, the FDA
Center for Food Safety and Applied Nutrition (CFSAN) concluded that the
FLAVR SAVR tomato was as safe and nutritious as other commonly consumed
tomatoes and ''that the use of the kan(r) gene and APH(3')IT will not
have a significant impact on the environment and that an environmental
impact statement is not required.'' On May 18, 1994, the FDA finished
its safety review of the FLAVR SAVR tomato and APH(3')II. The FDA
concluded that ''FLAVR SAVR(TM) tomatoes have not been significantly
altered when compared to varieties of tomatoes with a history of safe
use'' (Federal Register 59:26646) and ''that the use of aminoglycoside
3'phosphotransferase II is safe for use as a processing aid in the
development of new varieties of tomato, oilseed rape, and cotton
intended for food use'' (Federal Register 59:26700).

Document Number: 632

Recently it has been recognized that the action of the enzymes contained
in soybeans has negative effects on the qualities of soybean protein
foods. Based upon these findings, new manufacturing techniques for high
quality soybean protein foods have been developed through the control of
such enzyme action. For instance, new soybean seeds lacking
lipoxygenases have been developed which reduce the production of
grassy-beany flavors. The gel hardness of tofu can also be controlled
through the action of the lipoxygenases contained in soybeans. In
addition objectionable after-tastes develop through the hydrolysis of
soybean isoflavones by the beta-glucosidases which are found in
soybeans.
Genetic engineering of soybean proteins made a great progress in
improving their functional and nutritional qualities through the
modifications of genes encoding for soybean storage proteins. The
application of a bioreactor for soy sauce is also described as an
example of the progress on the biotechnology.

Document Number: 633

Identification of sources of resistance in cool season legumes to cyst
(Heterodera spp.), root-knot (Meloidogyne spp.), and stem nematode
(Ditylenchus dipsaci) is generally based on number of cysts on roots,
root-knot nematode induced gall index, and stem nematode reproduction in
shoot tissue, respectively. Various levels of resistance to cyst
nematodes have been detected in chickpea and pea. Resistance has also
been identified in chickpea, faba bean, and pea to the root-knot
nematodes. Broad based durable sources of resistance to plant parasitic
nematodes are required. Basic research is needed to develop transgenic
plants with resistance based on hatch stimulants, inhibitors, toxins, or
repellents found in antagonistic rhizosphere microorganisms. Selection
of genotypes that favor development of beneficial rhizosphere
microorganisms or root endophytes that increase the plant resistance to
nematode infection deserves attention.

Document Number: 634

Tomato fruit quality is determined by a complex set of characters. Fruit
used by the fresh market industry must have different attributes than
those used by the processing industries. We have determined the major
targets of the tomato industry and have isolated from the tomato itself
genes which based upon their biochemical function are expected to
improve texture and theology, colour, soluble solids and flavour. Our
research is aimed at proving this concept. So far we have demonstrated
that polygalacturonase (PG) is a major fruit quality gene providing the
processing tomato industry with improved viscosity. Commercial tomato
hybrids have been developed in collaboration with Petoseeds (USA) which
carry this effect gene. The regulatory system in Europe will be
reviewed. Field trials are covered by the EC directive 90/220, which has
now been implemented by all countries belonging to the European Union.
On the contrary there are no European-wide regulations in place to cover
the production of genetically modified plants as food sources. The
evolving food regulations will be reviewed and evaluated in the context
of competitiveness of the European plant biotechnology industry.

Document Number: 635

The potential of plant gene technology encompasses a multitude of
different techniques ranging from the isolation of useful genes, their
characterization and in vitro manipulation to the reintroduction of the
modified constructs into target plants, where they are expressed at a
rate that alters the phenotype of the plants. Genome analysis, on the
other hand, aims at characterizing the genome architecture and
function(s).
Plant gene technology has catalyzed progress in plant breeding, as will
be exemplified by a few examples, but has not yet been applied to food
legume improvement on a large scale. Genome analysis, however, has a
series of practical implications, as is illustrated by the successful
introduction of DNA fingerprint and PCR fingerprint techniques to
chickpea (Cicer arietinum L.) breeding and Ascochyta rabiei pathotyping.
The present overview addresses both areas of plant molecular biology to
illustrate their potential for food legume breeding.

Document Number: 636

A review of recent trends in animal science is used to provide a flavour
of how, from a purely technical viewpoint, evolving biotechnology may
influence animal production systems in the foreseeable future. The areas
considered are: the nutritive value of foods and forages, animal health
and welfare, the temporary alteration of production traits to meet an
immediate goal, and the permanent alteration of traits to meet an
immediate goal, and the permanent alteration of traits by genetic
selection or by the production of transgenic animals. In improving the
nutritive value Of foods, modern biotechnology has produced food grade
enzymes and amino acid supplements. Genetically engineered cereals,
grasses and legumes, rich in essential amino acids, are on the horizon.
In disease prevention, genetically engineered vaccines may be preferable
to conventional ones in that they have a defined composition, with a
degree of control during manufacture which allows amino acid
substitution within the peptide to confer specificity to a particular
serotype.
Modern techniques with the potential to promote a transitory alteration
in a production trait involve either immunization against an inhibitory
biochemical pathway or active promotion of a stimulatory pathway by the
administration of recombinant-derived hormones and proteins. Strategies
for permanent alteration of traits by gene transfer, initially directed
to enhancing growth, are now shifting in emphasis towards the transfer
of genes for enhanced nutrient digestion, disease resistance, and
improved product quality. The role of recombinant DNA technology in the
development of diagnostic probes for use in genetic selection programmes
and in the monitoring of disease is also expanding rapidly.
In all areas of animal science, molecular biology is promoting a better
understanding of the underlying mechanisms that control production. By
so doing it should ensure that science fulfils its primary role of
providing animal production industries with a range of options from
which to choose the most acceptable way forward.

Document Number: 637

Rapid strides have been made in the evolution of high-yielding varieties
in different tobaccos (Nicotiana spp) in India. Flue-cured virginia
tobacco showed an overall improvement of 90.6% for cured-leaf yield and
489.5% for bright-leaf yield during 1948-92. In bidi tobacco the yield
increased by 195.6% from 1950 to 1985. In chewing tobacco the yield
increased by 75% in Tamil Nadu (during 1948-89), 50%.in Bihar (during
1958-89) and 32% in Gujarat (during 1967-85). However, average yield in
a state in different tobaccos indicates that high productivity achieved
on research farms did not percolate to farmers' fields.
The major limiting factors in flue-cured virginia tobacco are frequent
drought spells, delayed monsoon resulting in late planting, incidence of
leaf-curl (Ruga tabaci Holmes) discase in Vertisols and southern
Alfisols of Andhra Pradesh, non-availability of varieties resistant to
root-knot nematode (Meloidogyne spp) (in northern Alfisols of Andhra
Pradesh) and aphid (Myzus spp) in Alfisols of Karnataka, and the
occurrence of broomrape (Orobanche cernua Leofl.). In bidi tobacco the
constraints are drought stress and biotic stresses of leaf-curl and
root-knot and the incidence of broomrape in Gujarat; aphid infestation
and incidence of black-shank [Phytophthora parasitica var nicotianae
(Breda de Haan) Tucker] as well as delayed and scanty rainfall in
Karnataka. In chewing tobacco the constraints are outbreak of
leaf-eating caterpillar (Spodoptera litura Fabricius), incidence of
tobacco mosaic (Marmor tabaci Holmes), delayed rain leading to late
planting in Tamil Nadu;  incidence of viral diseases in Bihar, root-knot
and leaf-curl diseases in Gujarat; brown-spot disease [Alternaria
alternata (Fr.) Keissler] in West Bengal and lack of high-yielding
varieties in Orissa. In natu tobacco the constraints are drought stress.
incidence of tobacco mosaic and outbreak of leaf-eating caterpillar. In
burley tobacco the constraints are, the incidence of black-shank and
frog-eye spot (Cercospora nicotianae Ellis & Everhart).
The approaches to overcome the constraints are to breed drought-tolerant
genotypes as well as to identify suitable hybrids that can maintain
stable yield under adverse conditions. The resistance breeding
programmes have to be pursued vigorously in view of the available donor
resistant genes for disease, viz tobacco mosaic: 'Vamorr 50';
black-shank: 'Bhavya', 'Line 1071' and N. plumbaginifolia Viviani;
root-knot: 'GT 5', 'Bhavya', N. amplexicaulis Burb. and N. longiflora
Cav.; and for pests, viz aphid: N. gossei Domin. and leaf-eating
caterpillar: 'DWFC' and Bacillius thuringiensis Berliner. For leaf-curl,
which involves the insect whitefly (Bemisia tabaci Genn.) and virus as
well as the complete-root parasite broomrape, no resistant genes are
available. Hence bio-technological means have to be adopted, such as
transgenic plants, protoplast fusion of interspecific hybrids, besides
continuing effort to spot resistant genotypes from somaclonal variants
and mutant populations.
Many recently released varieties show good promise in yield and
resistance to different factors. In flue-cured virginia tobacco, to meet
the growing demand at home and abroad, the production is targetted to
increase fran 112 million kg in 1990 to 192 million kg in 1995 from
Andhra Pradesh and Karnataka. The recently released high-yielding
varieties 'Gautami' and 'Virginia Tobacco 1158' (resistant to tobacco
mosaic) in Andhra Pradesh, and 'Bhavya' (resistant to black-shank,
tolerant to root-knot) and 'Swama' [resistant to powdery mildew
(Erysiphe cichoracearum DC.)] in Karnataka, show good potential of high
yield, In bidi tobacco, in the prevailing conditions of fluctuation in
area and production (due to inconsistent market trends and rainfall
pattern) as well as increase in demand, the high-yielding varieties 'GT
5' (tolerant to root-knot, high nicotine content of 8.62-9.00%, smooth
smoke generating 30% less smoke toxicants) and 'GT 7' (drought-tolerant)
show good potential in Gujarat. In chewing tobacco, with uncertain
rainfall, cost competition with other crops and lack of irrigation
sources, 'Meenakshi'- the recently released high-yielding variety -
shows good potential in Tamil Nadu for the small and scattered areas. It
also fits well in the most effective maize (Zea mays L)-tobacco-maize
cropping system in Bihar, where 'Sona' and 'Pusa Tobacco 76' show good
potential, whereas in Gujarat the high-yielding varieties 'GT 6' and 'GC
1'-with an ability to succeed the principal food crop, rice (Oryza
sativa L.)-show good potential. In natu tobacco the recently released
variety 'Natu Special' (>25-30% yield compared with 'WAF') has good
potential in cigarette natu tobacco-growing districts of Kurnool and
Mahaboobnagar of Andhra Pradesh.
The decennial tobacco productivity in India showed a progressive trend,
with an increase of 58% for all tobaccos, 37% for flue-cured virginia
and 66% for non-virginia tobacco from 1960-61 to 1990-91. Export of
tobacco and tobacco products from India increased by 68% in quantity and
709% in value from 1970-71 to 1990-91. With present consumption trend of
tobacco in different forms the world over, the tobacco export from India
is likely to increase.
Research is needed for breeding less health-hazardous tobaccos breeding
tobaccos for abiotic and biotic stresses, utilization of heterosis for
achieving stable yield under adverse conditions, breeding
low-photorespiratory tobaccos, breeding genotypes with high potassium
content for good quality, studies on leaf-surface chemistry for trichome
exudates that influence resistance to pests and on organoleptic
properties, and genetic tailoring of tobacco plants for alternative
uses, besides its narcotic value.

Document Number: 638

Glycinin is one of the dominant storage proteins of soybean seeds.
Previous studies showed that modified proglycinins, the variable regions
of which were inserted with oligopeptides composed of four continuous
methionines, could be synthesized in Escherichia coli and exhibited
better food functional properties than the native glycinin (C.-S. Kim,
S. Kamiya, T. Sato, S. Utsumi and M. Kito, Protein Engineering 3 (1990)
725-731). To investigate whether the modified glycinins would accumulate
in plant tissues, the normal and modified cDNAs were placed under the
control of the cauliflower mosaic virus 35S promoter in the binary
vector pBI121, and then inserted into the genome of tobacco by
Agrobacteriwn-mediated transformation. Both normal and modified
glycinins accumulated in the leaves, stems and seeds, and no differences
in the level of accumulation were observed among the expressed proteins
in leaves and seeds. The normal and modified glycinins expressed in each
tissue were present as the correct-size proteins. The proteins expressed
in each tissue were processed to the mature form, and those expressed in
the seeds assembled into hexamers. These results support the possibility
that we can create novel soybean plants which can produce modified
glycinins having better nutritional and functional properties.

Document Number: 639

Over the last 5000 years cereals have been bred for food, feed, and
beverages by selection of spontaneous mutations and random hybrids.
Since the turn of the century, crosses with defined parents, and since
1927 artificially induced mutations, have been used to create
variability on which selection of new varieties is based. It is pointed
out that hybrid corn and transfer of rust-resistant genes from wild
species into chromosomes of bread wheat was preceded by decades of basic
research. Genetic transformation is an additional tool for the breeder
to introduce novel genes in a rational manner and will complement but
not replace the existing efficient breeding methods. Genetic
transformation has been demonstrated in maize, rice, and wheat, while
techniques to obtain transgenic barley plants are still being developed.
Our present knowledge on the endosperm-specific expression of storage
proteins and the modulation of this expression by transcriptional
activators is reviewed. Breeding strategies for altered protein quality
and for proanthocyanidin-free malting barley are presented. Engineering
of an improved malt enzyme, a heat stable (1-3,1-4)-beta-glucanase, is
described. The enzyme is expected to survive, like alpha-amylases, the
kilning process and has been shown to act efficiently in the mashing
process for the elimination of water-soluble beta-glucans which impede
filtration of wort. The engineered enzyme is expressed in transformed
aleurone protoplasts and secreted from these cells and thus shown to be
operational in the tissue, where it is expected to work.
Hormone-regulated promoters for the expression of genes acting during
grain development and malting have been characterized. Prospects for the
production of polyhydroxyalkanoates and cyclodextrins in cereal grains
are discussed.

Document Number: 640

Recent progress in the field of biotechnology and the production of
transgenic livestock has raised a question regarding the need for the
regulation of these animals. There is also the need to regulate
nontransgenic animals resulting from transgenic animal research. It is
anticipated that several governmental agencies will be involved in
regulatory issues pertaining to these animals. The United States
Department of Agriculture (USDA), Food Safety and Inspection Service
(FSIS) will ultimately be responsible for ensuring that transgenic
animals intended for human consumption are wholesome, unadulterated, and
properly labeled. The FSIS has implemented a program for the regulation
of slaughtering nontransgenic animals resulting from transgenic animal
experiments. However, the FSIS has not yet approved any transgenic
livestock for slaughter. Scientists from the FSIS, in conjunction with
other government agencies, are currently developing guidelines for the
slaughter of transgenic animals.

Document Number: 641

The genetic events associated with traditional selection have
implications for the food safety of transgenic animals. Selection has
been empirical, relying on the use of the best animals for breeding.
Molecular techniques are now being used to identify the genes selected
and to describe the differences between alleles that are important in
selection to improve quantitative traits. The results of such analyses
provide background details of the genetic and physiological effects of
the traditional selection of animal lines. Examples of the kinds of
genes that may be subject to selection are those coding for peptide
hormones, steroid metabolic enzymes, the calcium-channel gating protein,
and genes of the major histocompatibility complex. Unselected genes,
sometimes with undesirable alleles, may be carried along as
''hitchhikers'' if they are closely linked to the selected gene. In
spite of this potential for physiologically dangerous genetic changes in
selected animals, hereditary food toxicity has never been associated
with a selected line of the common food animals. This is probably
because the allowable physiological range of results of selection is
limited by the requirement for healthy, productive animals. Based on
these limitations, foods from healthy transgenic animals produced for
the purpose of herd improvement are likely to be as safe as the foods
from the untransformed parental line. Animals are important indicators
of their own food safety.

Document Number: 642

Significant progress has been made in development of transgenic plants
containing agriculturally useful genes. Concurrent with scientific
advances has been development of a regulatory infrastructure within the
U.S. Department of Agriculture (USDA) for assessing safety of controlled
release of genetically engineered plants into the environment, as well
as creation of a food policy by the Food and Drug Administration (FDA).
Field trials and safety assessments of tomato containing an antisense
polygalacturonase gene (FLAVR SAVR(TM) tomato) have been conduced. A
detailed safety analysis of the kan(r) selectable marker was also done.
Based on these data plus nutritional measurements, lack of changes in
levels of natural toxins, and lack of any unintended changes, we have
requested that the USDA and FDA determine that this genetically
engineered tomato is safe for release into the environment and human
consumption.

Document Number: 643

The United Nations Conference on Environment and Development alerted the
world to the hazards presumptive if the planet's natural resources of
land, water, air, energy and biological organisms are not protected and
utilised more conservatively. UNCED declared two urgent research
priorities: global climate change and genetic diversity. Excessive use
of fossil fuels and a resultant atmospheric pollution forebodes higher
temperatures at the earth's surface. The consequences for agriculture
are unpredictable. Alternative sources of energy are described: electric
motors with high-capacity batteries and high energy fly-wheels;
solar-thermal and photovoltaic systems of power generation. Global
population will rise to at least 8 billion (1 billion = 10(9)) by 2025,
a rate of growth demanding an annual increase of 3% in agricultural
production. The principles of sustainable production in relation to land
and water management, genetic diversity among crop species, and food
science and technology are discussed. Among the many thousand crop
genotypes in the international germ plasm banks, more is known about
agronomic characters than about functional properties relevant to food
processing. Transgenic modifications broaden the spectrum of pest
resistance in crops, but too little is known of the nutritional
implications. Biotechnological advances, demands for non-polluting waste
disposal, consumer concerns for food safety, the need to provide food
security for a global population changing in age and income
distribution, offer exciting challenges to all food scientists and
technologists.

Document Number: 644

Progress in the development of gent-technology methods has enabled the
targeted transfer of genes between unrelated organisms. The results open
the way to a multitude of useful applications in very different sectors
of the economy. Here a review of applications and international
developments in gene technology as applied to nutrition and food
production is presented. Emphasis is placed on four main topics: 1)
Synthesis of single substances (e.g. enzymes. hormones, additives) by
genetically modified organisms, 2) The use of genetically engineered
microorganisms as starter cultures in food processing, 3) Transgenic
plants with newly introduced resistance characteristics or improved
storage or transportation properties. 4) Transgenic animals.
In the cases under study a risk assessment was carried out. Legal
regulations for foods derived from modern biotechnology and the proposal
for the ''Novel-Food'' regulation are described.

Document Number: 645

The regulation of transgenic plants is at the very early stages of
dialog between the U.S. Environmental Protection Agency (EPA), the Food
and Drug Administration (FDA), and the U.S. Department of Agriculture
(USDA). Herbicide resistance is discussed in the broader terms of
resistance management and of the cooperative efforts underway involving
governments, industry, academia, and the environmental community on an
international level. A new body, the International Organization for
Resistant Pest Management (IOPRM) was formed to implement pesticide
resistance management programs worldwide. The research base for
integrated pest management programs which form the context for pesticide
resistance management programs is in need of expansion. Public and
private efforts to expand this base converge in an ambitious
collaborative program between the U.S. EPA and USDA entitled the
National Integrated Pest Management Forum. Finally, the potential role
of EPA in the regulation of resistance management is discussed.

Document Number: 646

The application of recombinant DNA and related techniques to plants
opened up the potential to improve agronomic characters, food processing
traits and food quality properties of plants as food sources. This
review focuses on molecular approaches to improving the protein quality
and reducing flatulence in beans. Much is now known about the regulation
of seed-specific expression and organization of seed storage protein
genes. Several attempts have been made to increase the methionine
content of tobacco seeds using heterologous seed protein genes from
peas, common bean and soybean. Such techniques could also be used to
modify lysine, tryptophan and threonine contents in cereals and legumes.
Our knowledge of the biosynthetic pathways for raffinose
oligosaccharides allows addressing the problem of flatulence in bean
consumption and elucidating the role of these oligosaccharides in plant
biology through molecular techniques.
With the rapid progress being made in gene cloning and transformation
methods in plants, our knowledge of basic plant metabolic processes
could be the limiting factor in improving plants as food sources through
molecular strategies.

Document Number: 647

The author discusses factors which with the release of transgenic
animals could become a risk for the environment, the genetic
manipulation by means of viral vectors being of primary concern in this
discussion.  Health problems of transgenic animals and their impact on
the environment must also be taken into consideration as well as their
utilization for human food manufacture.

Document Number: 648

At present an enormous amount of chemicals is used in agriculture to
ensure high yields of all types of crops.  These chemicals are utilized
for various purposes, like fertilization or soil disinfection.  These
chemicals have excellently performed their function, but on the other
hand they have often a negative effect on the environment and sometimes
on food products, made from chemically treated crops.  Therefore certain
raw materials from plant origin have to be manufactured in a special way
to remove residual amounts of these chemicals.
The big challenge for the agriculture in the future will be how to
maintain the good agronomy of the present cultivation methods and to
diminish the amount of chemicals used.  Biotechnology may offer ways to
achieve this.  Specially selected or genetically modified bacteria can
play a role in biological fertilization.  Also modern biotechnology
offers the possibility to introduce genetic information encoding
resistance traits to certain pests.  The transfer of genes across
boarders between species is extremely useful as during evolution many
strategies to control competition between living organisms have been
generated.  Some of the possibilities to introduce pest resistance by
biotechnological approaches will be discussed.

