1: Curr Opin Biotechnol. 2009 Mar 9. [Epub ahead of print]



Transgenic plants for enhanced phytoremediation of toxic explosives.



Van Aken B.



Department of Civil and Environmental Engineering, West Virginia University,

Morgantown, WV 26506, USA.



Phytoremediation of organic pollutants, such as explosives, is often a slow and

incomplete process, potentially leading to the accumulation of toxic metabolites 

that can be further introduced into the food chain. During the past decade,

plants have been genetically modified to overcome the inherent limitations of

plant detoxification capabilities, following a strategy similar to the

development of transgenic crop. Bacterial genes encoding enzymes involved in the 

breakdown of explosives, such as nitroreductase and cytochrome P450, have been

introduced in higher plants, resulting in significant enhancement of plant

tolerance, uptake, and detoxification performances. Transgenic plants exhibiting 

biodegradation capabilities of microorganisms bring the promise of an efficient

and environmental-friendly technology for cleaning up polluted soils.



PMID: 19278849 [PubMed - as supplied by publisher]



2: J Biosci Bioeng. 2009 Mar;107(3):339-43.



Dynamics of genetically modified Lactococcus lactis in simulated natural

environments and impacts on microbial communities.



Hagi T, Minagawa A, Shima J.



National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642,

Japan.



The survival, impacts on microbiota and gene transfer ability of genetically

modified (GM) Lactococcus lactis containing ermAM gene in soil and water

environments were investigated. After inoculation of GM L. lactis into

environments, its DNA remained longer, and the microbial community was changed.

No transfer of chromosomal ermAM was observed.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19269603 [PubMed - in process]



3: J Agric Food Chem. 2009 Mar 5. [Epub ahead of print]



Simultaneous Detection of Recombinant DNA Segments Introduced into Genetically

Modified Crops with Multiplex Ligase Chain Reaction Coupled with Multiplex

Polymerase Chain Reaction.



Mano J, Oguchi T, Akiyama H, Teshima R, Hino A, Furui S, Kitta K.



National Food Research Institute, 2-1-12, Kannondai, Tsukuba, Ibaraki 305-8642,

Japan, and National Institute of Health Sciences, 1-18-1, Kamiyoga, Setagaya-ku, 

Tokyo 158-8501, Japan.



We developed a multiplex polymerase chain reaction (PCR)-multiplex ligase chain

reaction (LCR) (MPCR-MLCR) technique as a novel approach for the simultaneous

detection of recombinant DNA segments (e.g., promoters, trait genes, and

terminators) of genetically modified (GM) crops. With this technique, target DNA 

regions were amplified by multiplex PCR, the PCR products were then subjected to 

multiplex LCR as template DNAs, and the LCR products were then analyzed by

polyacrylamide gel electrophoresis and subsequent fluorescent scanning. Seven

recombinant DNA segments commonly introduced into some GM crop lines were

selected as target DNA regions. In addition, another MPCR-MLCR system for the

simultaneous detection of three endogenous DNA segments was designed as a

positive control test. The specificity and sensitivity of the method were

examined. The method allowed us to detect GM crops comprehensively and is

expected to be utilized for efficient screening of GM crops into which any one of

the seven recombinant DNA segments have been introduced, and for profiling the

segments.



PMID: 19265381 [PubMed - as supplied by publisher]



4: Science. 2009 Feb 20;323(5917):1004-5.



Richard Richards profile. Making every drop count in the buildup to a blue

revolution.



Finkel E.



Publication Types: 

    Biography

    Historical Article

    News

    Portraits



Personal Name as Subject: 

    Richards R



PMID: 19229014 [PubMed - indexed for MEDLINE]



5: Pol J Vet Sci. 2008;11(4):411-4.



Current issues connected with usage of genetically modified crops in production

of feed and livestock feeding.



Kwiatek K, Mazur M, Sieradzki Z.



Department of Hygiene of Animal Feedingstuffs, National Veterinary Research

Institute in Pulawy, Al. PartyzantÓÃw 57, 24-100 PuÕ?awy, Poland.

kwiatekk@piwet.pulawy.pl



Progress, which is brought by new advances in modern molecular biology, allowed

interference in the genome of live organisms and gene manipulation. Introducing

new genes to the recipient organism enables to give them new features, absent

before. Continuous increase in the area of the biotech crops triggers continuous 

discussion about safety of genetically modified (GM) crops, including food and

feed derived from them. Important issue connected with cultivation of genetically

modified crops is a horizontal gene transfer and a bacterial antibiotic

resistance. Discussion about safety of GM crops concerns also food allergies

caused by eating genetically modified food. The problem of genetic modifications 

of GM crops used for livestock feeding is widely discussed, taking into account

Polish feed law.



PMID: 19227143 [PubMed - in process]



6: Anal Bioanal Chem. 2009 Mar;393(6-7):1629-38. Epub 2009 Feb 19.



Sensitive and highly specific quantitative real-time PCR and ELISA for recording 

a potential transfer of novel DNA and Cry1Ab protein from feed into bovine milk.



Guertler P, Paul V, Albrecht C, Meyer HH.



Physiology Weihenstephan, Technische UniversitÓ?t MÓÌnchen, Weihenstephaner Berg 3,

85350, Freising, Germany. patrick.guertler@wzw.tum.de



To address food safety concerns of the public regarding the potential transfer of

recombinant DNA (cry1Ab) and protein (Cry1Ab) into the milk of cows fed

genetically modified maize (MON810), a highly specific and sensitive quantitative

real-time PCR (qPCR) and an ELISA were developed for monitoring suspicious

presence of novel DNA and Cry1Ab protein in bovine milk. The developed assays

were validated according to the assay validation criteria specified in the

European Commission Decision 2002/657/EC. The detection limit and detection

capability of the qPCR and ELISA were 100 copies of cry1Ab microL(-1) milk and

0.4 ng mL(-1) Cry1Ab, respectively. Recovery rates of 84.9% (DNA) and 97%

(protein) and low (<15%) imprecision revealed the reliable and accurate

estimations. A specific qPCR amplification and use of a specific antibody in

ELISA ascertained the high specificity of the assays. Using these assays for 90

milk samples collected from cows fed either transgenic (n = 8) or non-transgenic 

(n = 7) rations for 6 months, neither cry1Ab nor Cry1Ab protein were detected in 

any analyzed sample at the assay detection limits.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19225766 [PubMed - in process]



7: J Agric Food Chem. 2009 Feb 13. [Epub ahead of print]



Safety Assessment and Detection Method of Genetically Modified Chinese Kale (

Brassica oleracea cv. alboglabra ).



Lin CH, Lu CT, Lin HT, Pan TM.



Institute of Microbiology and Biochemistry, College of Life Science, National

Taiwan University, Number 1, Section 4, Roosevelt Road, Taipei 10617, Taiwan, and

Department of Food Science, Nutrition, and Nutraceutical Biotechnology, Shih

Chien University, Number 70, Ta-Chih Street, Taipei 10462, Taiwan.



Sporamins are tuberous storage proteins and account for 80% of soluble protein in

sweet potato tubers with trypsin-inhibitory activity. The expression of sporamin 

protein in transgenic Chinese kale (line BoA 3-1) conferred insecticidal activity

toward corn earworm [ Helicoverpa armigera (Hubner)] in a previous report. In

this study, we present a preliminary safety assessment of transgenic Chinese kale

BoA 3-1. Bioinformatic and simulated gastric fluid (SGF) analyses were performed 

to evaluate the allergenicity of sporamin protein. The substantial equivalence

between transgenic Chinese kale and its wild-type host has been demonstrated by

the comparison of important constituents. A reliable real-time polymerase chain

reaction (PCR) detection method was also developed to control sample quality.

Despite the results of most evaluations in this study being negative, the safety 

of sporamin in transgenic Chinese kale BoA 3-1 was uncluded because of the

allergenic risk revealed by bioinformatic analysis.



PMID: 19216530 [PubMed - as supplied by publisher]



8: Mol Ecol. 2009 Feb;18(4):569-71.



Comment on:

    Mol Ecol. 2009 Feb;18(4):750-61.



Unwanted transgenes re-discovered in Oaxacan maize.



Snow A.



Department of Evolution, Ecology, and Organismal Biology, Ohio State University, 

Columbus, OH 43210, USA. snow.1@osu.edu



Publication Types: 

    Comment



PMID: 19215581 [PubMed - indexed for MEDLINE]



9: Genes Dev. 2009 Feb 1;23(3):373-84.



Integration of transport-based models for phyllotaxis and midvein formation.



Bayer EM, Smith RS, Mandel T, Nakayama N, Sauer M, Prusinkiewicz P, Kuhlemeier C.



Institute of Plant Sciences, University of Bern, Bern, Switzerland.



The plant hormone auxin mediates developmental patterning by a mechanism that is 

based on active transport. In the shoot apical meristem, auxin gradients are

thought to be set up through a feedback loop between auxin and the activity and

polar localization of its transporter, the PIN1 protein. Two distinct molecular

mechanisms for the subcellular polarization of PIN1 have been proposed. For leaf 

positioning (phyllotaxis), an "up-the-gradient" PIN1 polarization mechanism has

been proposed, whereas the formation of vascular strands is thought to proceed by

"with-the-flux" PIN1 polarization. These patterning mechanisms intersect during

the initiation of the midvein, which raises the question of how two different

PIN1 polarization mechanisms may work together. Our detailed analysis of PIN1

polarization during midvein initiation suggests that both mechanisms for PIN1

polarization operate simultaneously. Computer simulations of the resulting dual

polarization model are able to reproduce the dynamics of observed PIN1

localization. In addition, the appearance of high auxin concentration in our

simulations throughout the initiation of the midvein is consistent with

experimental observation and offers an explanation for a long-standing criticism 

of the canalization hypothesis; namely, how both high flux and high concentration

can occur simultaneously in emerging veins.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19204121 [PubMed - indexed for MEDLINE]



10: Methods Mol Biol. 2009;483:89-101.



Production of antibody fragments in Arabidopsis seeds.



Van Droogenbroeck B, De Wilde K, Depicker A.



Department Plant Systems Biology, Flanders Institute for Biotechnology (VIB),

Gent, Belgium.



Plants offer a number of attractive benefits over conventional mammalian or

bacterial cell culture systems for the production of valuable pharmaceutical and 

industrial proteins. Currently, antibodies and their derived fragments represent 

the largest and most important group of biotechnological products in clinical

trials. In particular, single-chain antibodies are an interesting class of

biopharmaceuticals because they are able to overcome specific problems associated

with full-length antibodies. Another valuable antibody format is the scFv-Fc:

fusion of the Fc domain to a single-chain variable fragment restores antibody

effector functions, allows purification, and mimics, despite being a

'single-gene' product, the bivalent properties of a full-length IgG. Although

many different plant-based production platforms have been evaluated for antibody 

production, seeds are especially attractive because, as natural storage organs,

they provide an optimal biochemical environment for the accumulation and

long-term storage of large amounts of functional proteins. This chapter describes

how to achieve high-level seed-specific expression of antibody fragments, how to 

select the best transgenic lines, and how to evaluate the accumulation level in

the seed stocks from the selected lines.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19183895 [PubMed - indexed for MEDLINE]



11: Methods Mol Biol. 2009;483:69-87.



Production and localization of recombinant pharmaceuticals in transgenic seeds.



Rademacher T, Arcalis E, Stoger E.



Institute for Molecular Biology, RWTH Aachen, Worringerweg, Aachen, Germany.



Among the many plant-based production systems that have been developed for

pharmaceutical proteins, seeds have the useful advantage of accumulating proteins

in a relatively small volume, and recombinant proteins are very stable in dry

seeds allowing long-term storage and facilitating distribution before

processing.To take full advantage of the natural ability of endosperm cells to

store large amounts of protein in a protected subcellular environment, it is

useful to target recombinant proteins to appropriate storage organelles. In this 

chapter, we describe the distinct types of protein storage organelles in the

cereal endosperm and a protocol for the detection of recombinant proteins in

these organelles by immunofluorescence and immunogold labelling.The use of food

and feed crops for the production of pharmaceutical proteins such as edible

vaccines implies the need for strict separation of the transgenic seeds from the 

food and feed chain. For improved traceability visual markers may be co-expressed

with the gene of interest in engineered seeds. DsRed is one example for a

fluorescent protein that can be detected with high sensitivity using low tech

equipment. We therefore describe the generation of transgenic maize plants

expressing DsRed in a constitutive manner, and we point out the advantages of

using this marker during the process of transformation and selection of plant

tissue and later during breeding of transgenic lines into elite germplasm.



PMID: 19183894 [PubMed - indexed for MEDLINE]



12: BMC Genomics. 2009 Jan 29;10:55.



An Ac/Ds-mediated gene trap system for functional genomics in barley.



Lazarow K, LÓÌtticke S.



Biocenter Klein Flottbek, University of Hamburg, Hamburg, Germany.

lazarow@zedat.fu-berlin.de



BACKGROUND: Gene trapping is a powerful tool for gene discovery and functional

genomics in both animals and plants. Upon insertion of the gene trap construct

into an expressed gene, splice donor and acceptor sites facilitate the generation

of transcriptional fusions between the flanking sequence and the reporter.

Consequently, detection of reporter gene expression allows the identification of 

genes based on their expression pattern. Up to now rice is the only cereal crop

for which gene trap approaches exist. In this study we describe a gene trap

system in barley (Hordeum vulgare L.) based on the maize transposable elements

Ac/Ds. RESULTS: We generated gene trap barley lines by crossing Ac transposase

expressing plants with multiple independent transformants carrying the Ds based

gene trap construct GTDsB. Upstream of the beta-Glucuronidase start codon GTDsB

carries splice donor and acceptor sites optimized for monocotyledonous plants.

DNA blot analysis revealed GTDsB transposition frequencies of 11% and 26% in the 

F1 and F2 generation of gene trap lines and perpetuation of transposition

activity in later generations. Furthermore, analysis of sequences flanking

transposed GTDsB elements evidenced preferential insertion into expressed regions

of the barley genome. We screened leaves, nodes, immature florets, pollinated

florets, immature grains and seedlings of F2 plants and detected GUS expression

in 51% (72/141) of the plants. Thus, reporter gene expression was found in 24 of 

the 28 F1 lines tested and in progeny of all GTDsB parental lines. CONCLUSION:

Due to the frequent transposition of GTDsB and the efficient expression of the

GUS reporter gene, we conclude that this Ac/Ds-based gene trap system is an

applicable approach for gene discovery in barley. The successful introduction of 

a gene trap construct optimized for monocots in barley contributes a novel

functional genomics tool for this cereal crop.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19178688 [PubMed - indexed for MEDLINE]



13: Mod Healthc. 2009 Jan 12;39(2):17.



Battling the clones. CHW wants to avoid genetically altered foods.



Rhea S.



Publication Types: 

    News



PMID: 19172933 [PubMed - indexed for MEDLINE]



14: J Integr Plant Biol. 2009 Jan;51(1):58-66.



W55a encodes a novel protein kinase that is involved in multiple stress

responses.



Xu ZS, Liu L, Ni ZY, Liu P, Chen M, Li LC, Chen YF, Ma YZ.



National Key Facility of Crop Gene Resources and Genetic Improvement, Key

Laboratory of Crop Genetics and Breeding, Ministry of Agriculture, Institute of

Crop Science, the Chinese Academy of Agricultural Sciences, Beijing 100081,

China.



Protein kinases play crucial roles in response to external environment stress

signals. A putative protein kinase, W55a, belonging to SNF1-related protein

kinase 2 (SnRK2) subfamily, was isolated from a cDNA library of drought-treated

wheat seedlings. The entire length of W55a was obtained using rapid amplification

of 5' cDNA ends (5'-RACE) and reverse transcription-polymerase chain

reaction(RT-PCR). It contains a 1,029 -bp open reading frame (ORF) encoding 342

amino acids. The deduced amino acid sequence of W55a had eleven conserved

catalytic subdomains and one Ser/Thr protein kinase active-site that characterize

Ser/Thr protein kinases. Phylogenetic analysis showed that W55a was 90.38%

homologous with rice SAPK1, a member of the SnRK2 family. Using

nullisomic-tetrasomic and ditelocentric lines of Chinese Spring, W55a was located

on chromosome 2BS. Expression pattern analysis revealed that W55a was upregulated

by drought and salt, exogenous abscisic acid, salicylic acid, ethylene and methyl

jasmonate, but was not responsive to cold stress. In addition, W55a transcripts

were abundant in leaves, but not in roots or stems, under environmental stresses.

Transgenic Arabidopsis plants overexpressing W55a exhibited higher tolerance to

drought. Based on these findings, W55a encodes a novel dehydration-responsive

protein kinase that is involved in multiple stress signal transductions.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19166495 [PubMed - indexed for MEDLINE]



15: J Integr Plant Biol. 2009 Jan;51(1):35-44.



Ectopic overexpression of wheat adenosine diphosphate-ribosylation factor, TaARF,

increases growth rate in Arabidopsis.



Yao Y, Ni Z, Du J, Han Z, Chen Y, Zhang Q, Sun Q.



Department of Plant Genetics & Breeding and State Key Laboratory for

Agrobiotechnology, Key Laboratory of Crop Heterosis and Utilization, Ministry of 

Education, China Agricultural University, Beijing, China.



Differential gene expression between hybrids and their parents is considered to

be associated with heterosis. However, the physiological functions and possible

contribution to heterosis of these differentially expressed genes are unknown. We

have isolated one hybrid upregulated gene encoding putative wheat

ADP-ribosylation factor, designated TaARF. In this study, real-time quantitative 

reverse transcription-polymerase chain reaction analysis indicated that the TaARF

transcript was preferentially expressed in root, node and crown, and the

accumulation of TaARF mRNA in hybrid was more than 1.5-fold higher than that in

two parents. In order to understand possible roles of the putative wheat ARF

gene, TaARF was overexpressed in Arabidopsis, and the transgenic plants were

characterized. We show that ectopic overexpression of TaARF in Arabidopsis leads 

to increased leaf area, increased growth rate and earlier transition to

flowering, suggesting that TaARF plays significant roles in growth and

development. This study provides evidence demonstrating that TaARF plays

important roles in growth and development and we speculate that the upregulated

expression of this gene might contribute to the heterosis observed in wheat root 

and leaf growth.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19166492 [PubMed - indexed for MEDLINE]



16: J Dairy Sci. 2009 Feb;92(2):444-57.



Fate of lysostaphin in milk from individual cows through pasteurization and

cheesemaking.



Van Hekken DL, Wall RJ, Somkuti GA, Powell MA, Tunick MH, Tomasula PM.



Dairy Processing and Products Research Unit, USDA, Agricultural Research Service,

Eastern Regional Research Center, Wyndmoor, PA 19038, USA.

diane.vanhekken@ars.usda.gov



Transgenic cows secreting over 3 microg 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 degrees 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 degrees 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.



PMID: 19164654 [PubMed - indexed for MEDLINE]



17: J Genet Genomics. 2009 Jan;36(1):41-9.



Index selection on seed traits under direct, cytoplasmic and maternal effects in 

multiple environments.



Zhang W, Xu H, Zhu J.



Institute of Bioinformatics, College of Agriculture and Biotechnology, Zhejiang

University, Hangzhou 310029, China.



Crop seeds are important sources of protein, oil, and carbohydrates for food,

animal feeds, and industrial products. Recently, much attention has been paid to 

quality and functional properties of crop seeds. However, seed traits possess

some distinct genetic characteristics in comparison with plant traits, which

increase the difficulty of genetically improving these traits. In this study,

diallel analysis for seed models with genotype by environment interaction (GE)

effect was applied to estimate the variance-covariance components of seed traits.

Mixed linear model approaches were used to estimate the genetic covariances

between pair-wise seed and plant traits. The breeding values (BV) were divided

into two categories for the seed models. The first category of BV was defined as 

the combination of direct additive, cytoplasmic, and maternal additive effects,

which should be utilized for selecting stable cultivars over multi-environments. 

The three genetic effects, together with their GE interaction, were included in

the second category of BV for selecting special lines to be grown in specific

ecosystems. Accordingly, two types of selection indices for seed traits, i.e.,

general selection index and interaction selection index, were developed and

constructed on the first and the second category BV, respectively. These proposed

selection indices can be applied to solve the difficult task of simultaneously

improving multiple seed traits in various environments. Data of crop seeds with

regard to four seed traits and four yield traits based on the modified diallel

crosses in Upland cotton (Gossypium hirsutum L.) were used as an example for

demonstrating the proposed methodology.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19161944 [PubMed - in process]



18: Mol Plant Pathol. 2009 Jan;10(1):29-39.



Constitutive heterologous expression of avrXa27 in rice containing the R gene

Xa27 confers enhanced resistance to compatible Xanthomonas oryzae strains.



Tian D, Yin Z.



Temasek Life Sciences Laboratory, 1 Research Link, National University of

Singapore, Singapore 117604, Singapore.



The vascular pathogen Xanthomonas oryzae pv. oryzae (Xoo) and nonvascular

pathogen Xanthomonas oryzae pv. oryzicola (Xoc) cause bacterial blight (BB) and

bacterial leaf streak (BLS) diseases of rice, respectively. We have previously

identified the avirulence gene avrXa27 from Xoo PXO99(A), which specifically

induces the expression of the rice resistance gene Xa27, ultimately leading to

resistance against BB disease in rice. In this study, we have generated a

transgenic rice line (L24) that expresses avrXa27 constitutively under the

control of the PR1 promoter, and have examined its role in the host-pathogen

interaction. L24 is not more susceptible to BB, indicating that avrXa27 does not 

contribute to virulence. AvrXa27 retains avirulence activity in L24 and, after

crossing with a line containing Xa27, progeny display phenotypic changes

including inhibition of tillering, delay in flowering, stiff leaves, early leaf

senescence and activation of pathogenesis-related (PR) genes. On challenge with a

variety of compatible strains of Xoo and Xoc strain L8, lines with both avrXa27

and Xa27 also show enhanced resistance to bacterial infection. The induction of

Xa27 and subsequent inhibition of Xoc growth in Xa27 plants are observed on

inoculation with Xoc L8 harbouring avrXa27. Our results indicate that the

heterologous expression of avrXa27 in rice containing Xa27 triggers R

gene-specific resistance and, at the same time, confers enhanced resistance to

compatible strains of Xoo and Xoc. The expression of AvrXa27 and related proteins

in plants has the potential to generate broad resistance in plants.



Publication Types: 

    Research Support, Non-U.S. Gov't



PMID: 19161350 [PubMed - indexed for MEDLINE]



19: Rev Med Suisse. 2008 Dec 10;4(183):2709.



[Europe does not want to eat cloned food]



[Article in French]



Nau JY.



jynau@orange.fr



PMID: 19157291 [PubMed - indexed for MEDLINE]



20: Anal Chim Acta. 2009 Feb 16;634(1):75-82. Epub 2008 Dec 6.



Evaluation of stable isotope labelling strategies for the quantitation of CP4

EPSPS in genetically modified soya.



OcaÓÁa MF, Fraser PD, Patel RK, Halket JM, Bramley PM.



Centre for Chemical and Bioanalytical Sciences, Royal Holloway, University of

London, Egham TW20 0EX, UK.



The introduction of genetically modified (GM) crops into the market has raised a 

general alertness relating to the control and safety of foods. The applicability 

of protein separation hyphenated to mass spectrometry to identify the bacterial

enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) protein expressed in GM

crops has been previously reported [M.F. Ocana, P.D. Fraser, R.K.P. Patel, J.M.

Halket, P.M. Bramley, Rapid Commun. Mass Spectrom. 21 (2007) 319.]. Herein, we

investigate the suitability of two strategies that employ heavy stable isotopes, 

i.e. AQUA and iTRAQ, to quantify different levels of CP4 EPSPS in up to four GM

preparations. Both quantification strategies showed potential to determine

whether the presence of GM material is above the limits established by the

European Union. The AQUA quantification procedure involved protein

solubilisation/fractionation and subsequent separation using SDS-PAGE. A segment 

of the gel in which the protein of interest was located was excised, the stable

isotope labeled peptide added at a known concentration and proteolytic digestion 

initiated. Following recovery of the peptides, on-line separation and detection

using LC-MS was carried out. A similar approach was used for the iTRAQ workflow

with the exception that proteins were digested in solution and generated tryptic 

peptides were chemically tagged. Both procedures demonstrated the potential for

quantitative detection at 0.5% (w/w) GM soya which is a level below the current

European Union's threshold for food-labelling. In this context, a comparison

between the two procedures is provided within the present study.



Publication Types: 

    Evaluation Studies

    Research Support, Non-U.S. Gov't



PMID: 19154813 [PubMed - indexed for MEDLINE]