A reliable protocol for transformation of Catharanthus roseus through Agrobacterium tumefaciens.

Proliferation of axillary shoot buds and multiple shoot formation in Catharanthus roseus was obtained in 96 % explants on MS medium (3 % sucrose) containing NAA + BA. 2,4-D induced callusing in both, the nodal as well as in leaf segments. Leaf-derived callus was used for transformation with Agrobacterium tumefaciens LBA4404/pBI-S1. Bacterial cell concentration, duration of co-cultivation and acetosyringone concentration influenced transformation efficiency. Under optimal co-cultivation conditions, 98 % of the explants showed GUS expression. PCR based amplification of the transformed and subsequently selected callus tissue indicated the presence of uidA, Gly I and nptII genes.