Studies on in vitro regeneration and genetic transformation in chilli capsicum annuum L

Abstract

In vitro regeneration and Agrobacterium-mediated genetic transformation was studied newlinein popular local cultivar of chilli viz., Byadagi Kaddi. Different explants (cotyledon, hypocotyls newlineand cotyledonary nodal region) were cultured on MS medium supplemented with various newlinelevels of BAP (2 12 mg/l) and TDZ (0.1-1.0 mg/l) for adventitious shoot induction. Frequency newlineof adventitious shoot induction was significantly more in cotyledonary nodal region explant newlinethan cotyledon and hypocotyls explants. Adventitious shoot induction response was high on newlinemedium with 0.5 mg/l TDZ. Addition of 1 mg/l of GA3 to this medium resulted in higher newlinefrequency of shoot elongation. The elongated shoots were rooted on MS medium with 0.5 newlinemg/l IAA and 0.1 mg/l BAP. newlineDifferent concentrations and combinations of growth regulators were tried to study newlinecallus induction and regeneration from cotyledon, hypocotyls and cotyledonary nodal region newlineexplants. The media combination of 0.5 mg/l 2, 4-D and 0.5 mg/l kinetin resulted in higher newlinequantity of callus. Elimination of 2, 4-D from this media induced somatic embryos. Complete newlineregeneration was also observed on the same media but with very low frequency (10%). newlineShoot apical meristems were co-cultivated with Agrobacterium strain EHA105 newlinecarrying pBinBt3 construct having cry1Ac gene. The explants pre-cultured for 48 hours, newlineinfected with agrobacterial suspension having 0.5 × 108 cells/ml for 10 min along with newlineacetosyringene at 200 and#956;M for 48 hours yielded the transformants under in vitro conditions. newlineAnalysis of the putative transformants for the presence and expression of cry gene revealed newlinethat the transformation efficiency was 0.5 per cent. In two step RT-PCR amplicon of 1000 bp newlinerevealed the expression of transformed gene. newline