Efficient octopine Ti plasmid-derived vectors for Agrobacterium-mediated gene transfer to plants

Abstract

A two-component cloning system to transfer foreign DNA into plants was derived from the octopine Ti plasmid pTiB6S3. pGV2260 is a non-oncogenic Ti plasmid from which the T-region is deleted and substituted by pBR322. pGV831 is a streptomycin-resistant pBR325 derivative that contains a kanamycin resistance marker gene for plant cells and a site for cloning foreign genes between the 25-bp border sequences of the octopine T-region. Conjugative transfer of pGV831 derivatives to Agrobacterium and cointegration by homologous recombination between the pBR322 sequences present on pGV831 and pGV2260, can be obtained in a single step. Strains carrying the resulting cointegrated plasmids transfer and integrate T-DNA into the genome of tobacco protoplasts, and transformed tobacco calli are readily selected as resistant to kanamycin. Intact plants containing the entire DNA region between the T-DNA borders have been regenerated from such clones. In view of these properties we present pGV831 and its derivatives as vectors for efficient integration of foreign genes into plants.