Development of a protocol for Agrobacterium mediated transformation of Brassica oleraceae L var botrytis cv Early kunwari

The development of protocol for genetic transformation of plants holds tremendous potential for genetic manipulation of crops and other applications. In the present study, attempts have been made to develop a simple, reliable and reproducible protocol for Agrobacterium tumefaciens mediated transformation of hypocotyl explants of Brassica oleraceae L var botrytis. Different time of preincubation (24, 48, 72 and 96 hours), infection (1, 2, 3, 4, 5, 7, 10 minutes) and co-cultivation (24, 48 and 72 hours) were tested to have a combination of these parameters. It was observed that the combination of 72 hours pre-incubation, 7 minutes infection and 72 hours co-cultivation showed maximum frequency of GUS expression. Whereas other combinations tested either did not show GUS expression or the frequency of GUS expressing explants decreased. Acetosyringone has been shown to enhance the frequency of transformation.