Single-copy T-DNAs integrated at different positions in the Arabidopsis genome display uniform and comparable beta-glucuronidase accumulation levels.
This study aimed at determining whether transgene expression variability is observed in single-copy T-DNA plants and whether it can be correlated with the T-DNA integration position. Among a population of 135 Arabidopsis thaliana transformants, selected on the basis of antibiotic resistance marker expression, 21 single-copy T-DNA transformants were identified and characterized. In 19 of these 21 lines, 35S-beta-glucuronidase transgene expression, measured in two subsequent generations, was similar. This observation means that the intra-transformant variability was as high as the inter-transformant variability. Integration into an intergenic or genic region, into an exon or intron, in sense or antisense orientation, did not result in differential transgene expression. Remarkably, single-copy transformants were not always the highest expressers, implying that low transgene expression is not always induced by multicopy transformants. In only 2 of the 21 single-copy plants was the transgene expression more than 20-fold lower. However, characteristics of the insertion position in one of these lines did not differ significantly when compared to high-expressing lines. In the remaining line, methylation of the transgene was clearly demonstrated. In conclusion, screening for single-copy T-DNA transformants greatly enriches for stable and high transgene expression, because the integration position is not a major determinant of transgene expression variability in Arabidopsis.
De Buck
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S
Department of Plant Systems Biology, Flanders Interuniversity Institute for Biotechnology (VIB), Ghent University, Technologiepark 927, 9052, Gent, Belgium.
Windels
P
P
De Loose
M
M
Depicker
A
A
D017360:Arabidopsis / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry*; D018076:DNA, Complementary / Q000378:metabolism; D018744:DNA, Plant; D005470:Fluorometry; D017343:Genes, Plant; D018745:Genome, Plant; D005966:Glucuronidase / Q000378:metabolism*; D008957:Models, Genetic; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D030821:Plants, Genetically Modified; D017434:Protein Structure, Tertiary; D019076:Transgenes
D004269:DNA, Bacterial; D018076:DNA, Complementary; D018744:DNA, Plant; D016376:Oligonucleotides, Antisense; C036483:T-DNA; D005966:Glucuronidase