Construction Of RNAi Vector For Three Sweet Potato Virus And Transformation Of Sweet Potato

Sweet potato virus disease is a important factor influencing yield and quality of sweet potato. It can cause 50% reduction, or even no harvest.There is no effective method can prevent sweet potato virus disease now. Shooting tip is used to get virus-free plantlets, but they can be infected again. We hope to obtain transgenic sweet potato resisiting three common virus(sweet potato feathery mottle virus, SPFMV; Sweet latent virus, SPLV; Sweet potato virus G, SPVG) by RNAi technology(1) Conservative sequence of CP gene of three kinds of sweet potato was selected to synthesis a 600bp fragment (SPFLG). Then it was cloned reversely and repetitively to both sides of intron of pBlue NiRIntron. After that the fragment of SPFLG-Intron-SPFLG was inserted in pROK219 between 35S promoter and NOS terminator. Finally the fragment of 35S-SPFLG-Intron-SPFLG-NOS was cloned to pBIN 19, constructing RNAi vector transcripting stem-loop structure. It has been identified by digestion and PCR already.(2) The content of three virus was measured, before and after injecting RNAi vector to leaves of sweet potato that be infected by virus. The results showed that three sweet potato virus had been inhibited in injected group comparing to control group. The effect was most significant after 15 days. It indicates that the RNAi vector can inhibit three sweet potato virus by transient expression.(3)Suspended lines of Lizixiang and Beijing553 were established, and regenerative plants were obtained from Lizixiang suspended line.(4) About 4,000 explants of Lizixiang were transformed by Agrobacterium-mediated transformation, and 120 resistant callus were obtained, but no regenerative plant was obtained; About 2,800 explants of Lizixiang were transformed by Gene-gun transformation, and 10 regenerative plants were obtained,5 of that were identified to be transgenic plants by PCR.