Abstract
Global, comparative gene expression analysis is potentially a very powerful tool in the safety assessment of transgenic plants since it allows for the detection of differences in gene expression patterns between a transgenic line and the mother variety. In the present study, we compared the gene expression profile in developing seeds of wild type wheat and wheat transformed for endosperm-specific expression of an Aspergillus fumigatus phytase. High-level expression of the phytase gene was ensured by codon modification towards the prevalent codon usage of wheat genes and by using the wheat 1DX5HMW glutenin promoter for driving transgene expression. A 9K wheat unigene cDNA microarray was produced from cDNA libraries prepared mainly from developing wheat seed. The arrays were hybridised to flourescently labelled cDNA prepared from developing seeds of the transgenic wheat line and the mother variety, Bobwhite, at three developmental stages. Comparisons and statistical analyses of the gene expression profiles of the transgenic line vs. that of the mother line revealed only slight differences at the three developmental stages. In the few cases where differential expression was indicated by the statistical analysis it was primarily genes that were strongly expressed over a shorter interval of seed development such as genes encoding storage proteins. Accordingly, we interpret these differences in gene expression levels to result from minor asynchrony in seed development between the transgenic line and the mother line. In support of this, real time PCR validation of results from selected genes at the late developmental stage could not confirm differential expression of these genes. We conclude that the expression of the codon-modified A.␣fumigatus phytase gene in the wheat seed had no significant effects on the overall gene expression patterns in the developing seed.
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References
R Alba Z Fei P Payton Y Liu SL Moore P Debbie J Cohn M D’Ascenzo JS Gordon JKC Rose G Martin SD Tanksley M Bouzayen MM Jahn J Giovannoni (2004) ArticleTitleESTs, cDNA microarrays, and gene expression profiling: tools for dissectin plant physiology and development Plant J 39 697–714 Occurrence Handle10.1111/j.1365-313X.2004.02178.x Occurrence Handle15315633 Occurrence Handle1:CAS:528:DC%2BD2cXos1ylt7w%3D
H Brinch-Pedersen G Galili S Knudsen PB Holm (1996) ArticleTitleEngineering of the aspartate family biosynthetic pathway in barley (Hordeum vulgare L.) by transformation with heterologous genes encoding feed-back-insensitive aspartate kinase and dihydropicolinate synthase Plant Mol Biol 32 611–620 Occurrence Handle10.1007/BF00020202 Occurrence Handle8980513 Occurrence Handle1:CAS:528:DyaK2sXjt1ek
H Brinch-Pedersen A Olesen SK Rasmussen PB Holm (2000) ArticleTitleGeneration of transgenic wheat (Triticum aestivum L.) for constitutive accumulation of an Aspergillus phytase Mol Breed 6 195–206 Occurrence Handle10.1023/A:1009690730620 Occurrence Handle1:CAS:528:DC%2BD3cXjt1Cku7c%3D
H Brinch-Pedersen LD Sørensen PB Holm (2002) ArticleTitleEngineering crop plants: getting a handle on phosphate Trends Plant Sci 7 118–125 Occurrence Handle10.1016/S1360-1385(01)02222-1 Occurrence Handle11906835 Occurrence Handle1:CAS:528:DC%2BD38Xit1ClsbY%3D
H Brinch-Pedersen F Hatzack LD Sorensen PB Holm (2003) ArticleTitleConcerted action of endogenous and heterologous phytase on phytic acid degradation in seed of transgenic wheat (Triticum aestivum L.) Transgenic Res 12 649–659 Occurrence Handle10.1023/B:TRAG.0000005113.38002.e1 Occurrence Handle14713194 Occurrence Handle1:CAS:528:DC%2BD3sXpt1Snu7k%3D
F Cellini A Chesson I Colquhoun A Constable HV Davies KH Engel AM Gatehouse S Kärenlampi EJ Kok JJ Leguay S Lehesranta HP Noteborn J Pedersen M Smith (2004) ArticleTitleUnintended effects and their detection in genetically modified crops Food Chem Toxicol 42 1089–1125 Occurrence Handle15123383 Occurrence Handle1:CAS:528:DC%2BD2cXjs1ertLk%3D
AH Christensen RA Sharrock PH Quail (1992) ArticleTitleMaize polyubiquitin genes: structure, thermal perturbation of expression and transcript splicing, and promoter activity following transfer to protoplasts by electroporation Plant Mol Biol 18 675–689 Occurrence Handle10.1007/BF00020010 Occurrence Handle1313711 Occurrence Handle1:CAS:528:DyaK3sXksVahtr8%3D
BC Clarke M Hobbs D Skylas R Appels (2000) ArticleTitleGenes active in developing wheat endosperm Funct Integr Genomics 1 44–55 Occurrence Handle10.1007/s101420000008 Occurrence Handle11793221 Occurrence Handle1:CAS:528:DC%2BD3cXktVGhtLg%3D
S Dudoit YH Yang MJ Callow TP Speed (2002) ArticleTitleStatistical methods for identifying differentially expressed genes in replicated cDNA microarray experiments Stat Sinica 12 111–140
MM Eisen PO Brown (1999) ArticleTitleDNA arrays for analysis of gene expression Method Enzymol 303 179–205 Occurrence Handle1:CAS:528:DyaK1MXltFOns7o%3D
O Fiehn S Kloska T Altman (2001) ArticleTitleIntegrated studies on plant biology using multiparallel techniques Curr Opin Biotechnol 12 82–86 Occurrence Handle10.1016/S0958-1669(00)00165-8 Occurrence Handle11167078 Occurrence Handle1:CAS:528:DC%2BD3MXhslGms7o%3D
EJ Kok HA Kuiper (2003) ArticleTitleComparative safety assessment for biotech crops Trends Biotechnol 21 439–444 Occurrence Handle10.1016/j.tibtech.2003.08.003 Occurrence Handle14512230 Occurrence Handle1:CAS:528:DC%2BD3sXnsVKnsrk%3D
HA Kuiper EJ Kok K-H Engel (2003) ArticleTitleExploitation of molecular profiling techniques for GM food safety assessment Curr Opin Biotechnol 14 238–243 Occurrence Handle10.1016/S0958-1669(03)00021-1 Occurrence Handle12732328 Occurrence Handle1:CAS:528:DC%2BD3sXjsVGksr4%3D
DJ Leader (2005) ArticleTitleTranscriptional analysis and functional genomics in wheat J Cereal Sci 41 149–163 Occurrence Handle10.1016/j.jcs.2004.10.006 Occurrence Handle1:CAS:528:DC%2BD2MXms1GnsA%3D%3D
P Lucca R Hurrell I Potrykus (2001) ArticleTitleGenetic engineering approaches to improve the bioavailability and the level of iron in rice grains Theor Appl Genet 102 392–397 Occurrence Handle10.1007/s001220051659 Occurrence Handle1:CAS:528:DC%2BD3MXisVynsb8%3D
SE Ouakfaoui B Miki (2005) ArticleTitleThe stability of the Arabidopsis transcriptome in transgenic plants expressing the marker genes nptII and uidA Plant J 41 791–800 Occurrence Handle10.1111/j.1365-313X.2005.02350.x Occurrence Handle15743445
L Pasamontes M Haiker M Wyss M Tessier APGM Loon Particlevan (1997) ArticleTitleGene cloning, purification, and characterization of a heat-stable phytase from fungus Aspergillus fumigatus Appl Environ Microbiol 63 1696–1700 Occurrence Handle9143104 Occurrence Handle1:CAS:528:DyaK2sXjtFGnsbo%3D
J Quackenbush (2002) ArticleTitleMicroarray data normalization and transformation Nat Biotechnol Suppl 22 496–501
JC Rogers C Milliman (1983) ArticleTitleIsolation and sequence analysis of a barley α-amylase cDNA clone J Biol Chem 258 8169–8174 Occurrence Handle6190808 Occurrence Handle1:CAS:528:DyaL3sXltVShur8%3D
PR Shewry F Beaudoin J Jenkins S Griffiths-Jones ENC Mills (2002) ArticleTitlePlant protein families and their relationships to food allergy Biochem Soc Trans 30 906–910 Occurrence Handle12440943 Occurrence Handle1:CAS:528:DC%2BD38XovF2isrY%3D
PR Shewry AS Tatham NG Halford JHA Barker U Hannappel P Gallois M Thomas M Kreis (1994) ArticleTitleOpportunities for manipulating the seed protein composition of wheat and barley in order to improve quality Transgenic Res 3 3–12 Occurrence Handle10.1007/BF01976021 Occurrence Handle8142950 Occurrence Handle1:CAS:528:DyaK2cXmsl2ju7c%3D
DH Simmonds TP O’Brien (1981) Morphological and biochemical development of the wheat endosperm Y Pomeranz (Eds) Advances in Cereal Science and Technology NumberInSeriesIV American Society of Cereal Chemists, Inc St. Paul, Minnesota 6–70
Smyth GK (2004) Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol 3(1)
GK Smyth TP Speed (2003) ArticleTitleNormalization of cDNA microarray data Methods 31 265–273 Occurrence Handle10.1016/S1046-2023(03)00155-5 Occurrence Handle14597310 Occurrence Handle1:CAS:528:DC%2BD3sXos1Chur4%3D
GK Smyth Y-H Yang TP Speed (2003) Statistical issues in microarray data analysis MJ Brownstein AB Khodursky (Eds) Functional Genomics: Methods and Protocols, Methods in Molecular Biology NumberInSeries224 Humana Press Totowa, NJ 111–136
CJ Thompson NR Novva R Tizard R Crameri JE Davies M Lauwereys J Botterman (1987) ArticleTitleCharacterization of the herbicide resistance gene bar from Streptomyces hygroscopicus EMBO J 6 2519–2523 Occurrence Handle1:CAS:528:DyaL2sXlsVygsr0%3D
RN Trethewey (2004) ArticleTitleMetabolite profiling as an aid to metabolic engineering in plants Curr Opin Plant Biol 7 196–201 Occurrence Handle10.1016/j.pbi.2003.12.003 Occurrence Handle15003221 Occurrence Handle1:CAS:528:DC%2BD2cXhslChsrs%3D
ID Wilson GLA Barker RW Beswick SK Shepherd C Lu JA Coghill D Edwards P Owen R Lyons JS Parker JR Lenton MJ Holdsworth PR Shewry KJ Edwards (2004) ArticleTitleA transcriptomics resource for wheat functional genomics Plant Biotechnol J 2 495–506
HY Yang S Dudoit P Luu DM Lin V Peng J Ngai TP Speed (2002) ArticleTitleNormalization for cDNA microarray data: a robust composite method addressing single and multiple slide systematic variation Nucleic Acids Res 30 e15 Occurrence Handle11842121
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Gregersen, P.L., Brinch-Pedersen, H. & Holm, P.B. A Microarray-Based Comparative Analysis of Gene Expression Profiles During Grain Development in Transgenic and Wild Type Wheat. Transgenic Res 14, 887–905 (2005). https://doi.org/10.1007/s11248-005-1526-y
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DOI: https://doi.org/10.1007/s11248-005-1526-y