Agrobacterium tumefaciens-mediated transformation of cauliflower, Brassicaoleracea var. botrytis

We have developed an efficient and simpler method for genetic transformatio n and regeneration of cauliflower, Brassica oleracea var. botrytis plants. Explants from 4-day old seedlings were inoculated and cocultivated with Agr obacterium tumefaciens strain LBA4404 harbouring a binary vee tor with the neomycin phosphotransferase-II gene under the regulatory control of nopalin e synthase promoter and terminator sequences, permitting transformed shoots to be selected on kanamycin containing medium. After three months rooted t ransformed plantlets were successfully transferred and grown under glasshou se conditions. Higher numbers of transformed plants were obtained from coty ledon than hypocotyl explants, presumably indicating cotyledons of cauliflo wer are more amenable to genetic transformation. Integration and expression of the introduced transgene were analysed by DNA gel blot and PCR analysis and NPT-II expression assay. Factors influencing transformation efficiency include explant age, concentration of bacterium used for infection, durati on of infection and cocultivation with Agrobacterium, Transgenic plants of three commercial genotypes of cauliflower were produced using this method. We also show that introduction of antisense Bcpl (pollen-specific gene) lin ked to a pollen-specific promoter (Lat52) resulted in the expected sterilit y of 50% pollen carrying this transgenic construct.