Genetic Transformation Of Plant Disease-resistance Gene Sncl In Parental Lines Of Super Hybrid Rice

Rice (Oryza sativa L.) is one of the most important grain crops in the world and is a kind of staple food for about 40% global population. Traditional genetic breeding techniques have made significant contributions to improving the quality and increasing the yield of rice. However, the genetic improvement of rice, especially the improvement of resistance to insects and diseases, has been severely challenged due to the environmental deterioration and the limitation of conventional breeding method. In this study, two parental lines of super hybrid rice, a light and temperature sensitive genie male sterile line P88S and a paternal line 0293 were used as materials to study the genetic transformation of the broad-spectrum disease-resistance gene sncl into the mature embryo callus of the two varieties by Agrobacterium-mediaXed transformation. After screening and differentiation of the callus, the new germplasms with higher resistance to pathogen are expected to be obtained for rice hybridation.The main results in the experiments are as the follows:1. The systems for callus induction of P88S and 0293 were established. The NMB induction medium which including N6 Macronutrient component+MS Micronutrient component+vitamin B5+3.0 mg/L 2-4,D+0.2 mg/L 6-BA+500 mg/L proline+500 mg/LCH+30 g/L sucrose+7-8 g/L agar (pH 5.8) is found to be the optimal medium for callus induction and subculture, the callus induction rates of both varieties are above 92%, and the subcultural calluses of the two varieties on NMB medium increase by 185%(up to 0.233 g) and 220%(up to 0.252 g), respectively.2. The Agrobacterium of EHA105 with the target gene of sncl was obtained through plasmid transformation. The Agrobacterium-mediated callus transformation system was optimized from four aspects of Agrobacterium cultivation mode, dipping method, drying time after dipping and co-culture time. LB liquid is chose as medium for Agrobacterium cultivation, oscillating for 15 min first, then vacuuming for 15 min, and then manual oscillating for 5 min for dipping of callus. After drying for 4 h and co-cultured for 2 d, the resistant callus obtained rate can reach to 29.78% under these optimum conditions.3. Through screening and regeneration of the disease-resistance callus, the transformed plants of P88S and 0293 which contained sncl gene is obtained and confirmed by PCR check.