ANALYSIS OF OCTOPINE LEFT BORDER-DIRECTED DNA TRANSFER FROM AGROBACTERIUM TO PLANTS

We constructed a binary plasmid, pVR30, with a neomycin phosphotransfe rase II (nptII) plant expression cassette flanked by a pTiA6 left bord er on its right and a pTiA6 right border on its left. This plasmid was used to study transfer of DNA to plants from a left border in the pre sence of a right border. Infection of tobacco leaf discs with a wild t ype octopine strain of Agrobacterium tumefaciens harbouring the binary plasmid resulted in the generation of kanamycin resistant calli at 18 to 26% frequency. Southern hybridization analysis of DNA isolated fro m eight transformed lines to different probes indicated that left bord er could mediate DNA transfer to plants in the presence of a right bor der in cis. Our results also suggest that transfer events correspondin g to transfer of T-centre DNA of octopine Ti plasmid pTiA6 do occur. W e have shown the relevance of left border-initiated T-DNA transfer by specifically selecting for such events and have confirmed it by Southe rn hybridization analysis. We also found that a border could be skippe d in a few T-DNA transsfer events.