A Microarray-Based Comparative Analysis of Gene Expression Profiles During Grain Development in Transgenic and Wild Type Wheat
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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.
Key wordscDNA microarrays gene expression phytase safety assessment transgenic wheat
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- Alba, R, Fei, Z, Payton, P, Liu, Y, Moore, SL, Debbie, P, Cohn, J, D’Ascenzo, M, Gordon, JS, Rose, JKC, Martin, G, Tanksley, SD, Bouzayen, M, Jahn, MM, Giovannoni, J 2004ESTs, cDNA microarrays, and gene expression profiling: tools for dissectin plant physiology and developmentPlant J39697714CrossRefPubMedGoogle Scholar
- Brinch-Pedersen, H, Galili, G, Knudsen, S, Holm, PB 1996Engineering of the aspartate family biosynthetic pathway in barley (Hordeum vulgare L.) by transformation with heterologous genes encoding feed-back-insensitive aspartate kinase and dihydropicolinate synthasePlant Mol Biol32611620CrossRefPubMedGoogle Scholar
- Dudoit, S, Yang, YH, Callow, MJ, Speed, TP 2002Statistical methods for identifying differentially expressed genes in replicated cDNA microarray experimentsStat Sinica12111140Google Scholar
- Eisen, MM, Brown, PO 1999DNA arrays for analysis of gene expressionMethod Enzymol303179205Google Scholar
- Quackenbush, J 2002Microarray data normalization and transformationNat Biotechnol Suppl22496501Google Scholar
- Simmonds, DH, O’Brien, TP 1981Morphological and biochemical development of the wheat endospermPomeranz, Y eds. Advances in Cereal Science and TechnologyAmerican Society of Cereal Chemists, IncSt. Paul, Minnesota670Google Scholar
- Smyth GK (2004) Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol 3(1)Google Scholar
- Smyth, GK, Yang, Y-H, Speed, TP 2003 Statistical issues in microarray data analysisBrownstein, MJKhodursky, AB eds. Functional Genomics: Methods and Protocols, Methods in Molecular BiologyHumana PressTotowa, NJ111136Google Scholar
- Thompson, CJ, Novva, NR, Tizard, R, Crameri, R, Davies, JE, Lauwereys, M, Botterman, J 1987Characterization of the herbicide resistance gene bar from Streptomyces hygroscopicusEMBO J625192523Google Scholar
- Wilson, ID, Barker, GLA, Beswick, RW, Shepherd, SK, Lu, C, Coghill, JA, Edwards, D, Owen, P, Lyons, R, Parker, JS, Lenton, JR, Holdsworth, MJ, Shewry, PR, Edwards, KJ 2004A transcriptomics resource for wheat functional genomicsPlant Biotechnol J2495506Google Scholar