The SEV System: A New Disarmed Ti Plasmid Vector System for Plant Transformation

Abstract

A derivative of the octopine-type Ti plasmid, pTiB6S3, was constructed in which the TL-DNA oncogenic functions, the TL-DNA right border sequence and all of TR-DNA were deleted and replaced with the kanamycin antibiotic resistance marker from Tn903 (601). The resulting avirulent plasmid, pTiB6S3–SE contains only the TL-DNA left border sequence and a region (1.6 kb) of homologous DNA (LIH) to allow recombination with intermediate vectors such as pMON120 or pMON200. Cointegrate formation between pTiB6S3–SE and pMON200, which contains a chimeric plant kanamycin resistance gene (Kmr), an intact nopaline synthase gene (NOS), a poly linker region to facilitate the insertion of foreign genes into the vector and a functional T-DNA right border sequence, results in the formation of a selectable, avirulent T-DNA. This vector system is referred to as the SEV system (Split End Vector) since the T-DNA border sequences are present on separate plasmids prior to recombination. Plant cells (tobacco, petunia and tomato) transformed with SEV T-DNA are easily identified by their resistance to kanamycin and their ability to synthesize nopaline. A major advantage of the new disarmed system is that all transformed colonies can potentially be regenerated into intact plants that normally express and segregate the inserted foreign DNA sequences.