Influence of genotype on Agrobacterium-mediated transformation of tomato

An efficient and reproducible regeneration protocol for tomato cultivars Arka Meghali, Arka Saurabh and Arka Vikas was developed using cotyledon and hypocotyl explants. Best regeneration response for all the three genotypes was obtained on MS medium supplemented with BAP (2 mg/l) and IAA (0.1 mg/l). All the three genotypes were transformed with Agrobacterium strain LBA 4404 containing the binary vector pCambia 2301 harbouring npt II gene as selectable marker and GUS as reporter gene. The transformed shoots were selected on regeneration medium containing kanamycin (75 mg/l) and cefotaxime (500 mg/l) and subsequently rooted in the presence of kanamycin (50 mg/l). Genotypic differences in the regeneration potential as well as in regeneration of putatively transformed plants were observed. Maximum percentage of transformed plants was obtained from genotype Arka Saurabh which exhibited the least regeneration potential while the genotype Arka Meghali with maximum regeneration potential gave rise to lower percentage of transformants. However, when transformation was expressed as transformation efficiency (number of putative transformants testing positive for reporter gene over the number of putative transformants) the trend remained same for the genotype Arka Saurabh which exhibited maximum transformation efficiency of 50% with least regeneration potential, while the trend was reversed for Arka Meghali the genotype with maximum regenerating ability recording maximum transformation efficiency of 50%. The presence of transgene was confirmed through PCR amplification of npt II gene as well as GUS assay. The results of the present study suggests that there may not always be a direct relationship between regeneration potential of the genotype and its transformation frequency or efficiency and it may vary with different genotypes.