Protocol for Direct Transformation through Gene Gun in Capsicum annuum L.Cv. Mathania
protocol for direct transformation through biolistic gun has been established for chilli (Capsicum annuum L. cv. Mathania). A disarmed strain of Agrobacterium tumefaciens EHA 105 carrying a binary vector plasmid p35SGUSINT has been used for transformation. This vector contains neomycin phosphotransferase gene (npt II), whose expression confirms Kanamycin resistance in transformants. In addition to npt II, plasmid encodes -glucuronidase, reporter enzyme used for studying the expression of foreign genes in plants. We report for the first time, the use of Nitrogen gas for the biolistic transformation of chilli. We found physical parameter like 6 cm target distance with 900 psi rupture disk for partical gun experiment the most efficient for transient GUS expression. In case of nitrogen gas the frequency of transient GUS expression was better both in leaves (67.74%) and hypocotyl (69.18%) and frequency of conversion of transient to stable transformation was 3.4% in leaves and 4.0% in hypotocyls as against 63.91 % transient GUS expression showed by leaves and 63.77% showed by hypocotyls when helium gas was used in biolistic transformation and stable transformation frequency was 3.0% in leaves and 4.0% in hypocotyls. The transgenic nature of the regenerated plants was confirmed by the histochemical staining of GUS, and polymerase chain reaction (PCR) analysis of npt II gene.