Studies On Construction Of Expression Vector With Resveratrol Synthase Gene From Peanut And Genetic Transformation System Of Cucumber

Stilbene synthase(STS)gene copied mostly from Arachis, Vitis, Pinus, includes resveratrol synthase (RS)gene and pinosylvin synthase(PS) gene. Progresses in the gene were summarized in this paper, and emphasis is the number, the structure and the homology of the gene. Inducing expression and regulatory of stilbene synthase gene were analyzed. And the resistance of transgenic plants, as tobacco, rice, wheat and alfalfa, were discussed.To further study the function of Resveratrol Synthase , RS gene was transferred to cucumber in this experiment. The RS gene has been already cloned in vectors pUCRSHl and pUCRSH2 before. pBI and pBI121 were used as expression vectors. Then the expression vectors and cloning vectors containing RS gene were digested by the suitable restriction enzymes, the fragments of interest were purified and ligated to each other by T4 DNA ligase. The construct was then transformed into a host strain and screened under the selective pressure of Kanamycin. It was confirmed that the recombinant expression plasmid pBI-RS pBI121-RS has been constructed successfully by examining them with PCR and restriction enzyme digestion.The recombinant expression plasmid was extracted and introduced to Agrobacterium tumefaciens strain LBA4404 with the method of freeze-melt. After culturing and screening the right transformants, the plasmid was extracted with methods of mini-Ti plasmid extracting. It was confirmed with PCR that the recombinant expression plasmidcontaining RS gene has been successfully introduced into Agrobacterium tumefaciens strain LBA4404.There are several factors that can affect the regeneration of adventitious bud: disinfection of cucumber seeds, selection of explant, hormone, differentiation of adventitious bud and resistance screening etc. The efficient protocol for transforming cucumber mediated by Agrobacterium tumefaciens has been established. The explant of cucumber was infected by Agrobacterium tumefaciens strain LBA4404 which contained pBI-RS, pBI121-RS plasmid and the resistance plants was obtained.