Effect of ploidy and homozygosity an transgene expression in primary tobacco transformants and their androgenetic progenies

Expression of a transgene is rarely analysed in the androgenetic progenies of the transgenic plants. Here, we report differential transgene expression in androgenetic haploid and doubled haploid (DH) tobacco plants as compare d to the diploid parental lines, thus demonstrating a gene dosage effect. U sing Agrobacterium-mediated transformation, and bacterial reporter genes en coding neomycin phosphotransferase (nptII) and beta-glucuronidase (uidA/ GU S), driven respectively by the mas 1' and mas 2' promoters, we have generat ed more than 150 independent transgenic (R-0) Nicotiana tabacum plants cont aining one or more T-DNA copies. Transgene analyses of these R-0, their sel fed R-1 lines and their corresponding haploid progenies showed an obvious p osition effect (site of T-DNA insertion on chromosome) on uidA expression. However, transgene (GUS) expression levels were not proportional to transge ne copy number. More than 150 haploids and doubled haploids, induced by tre atment with colchicine, were produced from 20 independent transgenic R-0 pl ants containing single and multiple copies of the uidA gene. We observed th at homozygous DH plants expressed GUS at approximately 2.9-fold the level o f the corresponding parental haploid plants. This increase in transgene exp ression may be attributed mainly to the increase (2-fold) in chromosome num ber. Based on this observation, we suggest a strong link between chromosome number (ploidy dosage effect) and transgene expression. In particular, we demonstrate the effect on its expression level of converting the transgene from the heterozygous (in R-0 plants) to the homozygous (DH) state: e.g. an increase of 50% was observed in the homozygous DH as compared to the origi nal heterozygous diploid plants. We propose that ploidy coupled with homozy gosity can result in a new type of gene activation, creating differences in gene expression patterns.