APX And Hevein Gene Transfer Research For Sisal

Sisal is the most important hemp cash crops in the tropical areas of China. Sisal and H.11648is the most popular variety, with high yield and cold resistance, but it is susceptibility Phytophtora nicotianae. Thus it is ugent need to breed new varieties with high disease resistance in sisal production.In order to cultivate new disease-resistant varieties, in this research of sisal, in this research, sisal H.11648callus induced from stem tips was used for testing. Various factors in Agrobacterium-mediated transformation. The results show that:adding50mg/L ascorbic acid to infection solution and co-cultivation medium solution, adding0.1%Tween in infection solution could significantly improve the conversion efficiency; Pre-incubation time2d, bacterial concentration OD600=0.5, infection time25min, co-culture4d at22℃, low concentrations of continuing screening methods could improve the efficiency of genetic transformation for sisal.Based on PCAMBIA3300plasmid, the APX’s plant expression vector Pcambia3300-35S-APX-nos driven by35s was successfully built and imported in sisal by agrobacterium EHA105. Enzyme validation showed that this recombinant expression vector has been transfer red into the agrobacterium EHA105.Based on using the above optimized system, APX gene was transferred into sisal callus. Total51resistant plants with APX single gene was obtained. PCR positive rate was37.25%,15plants with RT-PCR positive. Three of them were highly resistant plants and five were resistant to Phytophthora. APX genes showed a certain degree of resistance to Phytophthora. Thus, APX gene had been integrated into the genome of sisal.Based on PCAMBIA3300plasmid, the hevein’s plant expression vector PCAMBIA3300-hrs203J-hevein-nos driven by hrs230J was also successfully built.. Using the optimized system,30callus was infected by EHA105with Pcambia3300-hrs203J-hevein-nos,11seedlings with anti-PPT, and3plants with PCR positive, The positive rate was27.3%,3plants with RT-PCR positive were obtained. Phytophthora resistance test showed that three plants are highly resistant. In summary, Thus Hevein gene had been also integrated into the genome of sisal.