农杆菌介导的双SBE基因RNAi载体对水稻的遗传转化

Genetic Transformation with RNAi Vector of Two Starch Branching Enzyme on Oryza sativa L.via Agrobacterium

[目的]提高水稻中直链淀粉的含量,扩大稻米的利用范围。[方法]以水稻中花11品种为材料,通过农杆菌介导法,侵染水稻幼胚诱导出的愈伤组织,将淀粉分支酶基因SBE1和SBE3同时导入受体愈伤组织,经过一系列的共培养、预分化、分化、生根壮苗后,获得含有转基因的转化植株,并对转化植株进行PCR鉴定。[结果]通过用根癌农杆菌介导法,侵染由水稻幼胚诱导的274粒愈伤组织,经过共培养、筛选,得到177粒抗性愈伤,然后将抗性愈伤预分化、分化得到103株分化苗,生根壮苗后获得49株转基因植株;从49株转基因植株中随机提取34株的基因组DNA,利用PCR技术从基因组DNA中扩增得到了片段大小约为500 bp的潮霉素基因序列。[结论]初步证实转基因植株的真实性,淀粉分支酶基因SBE1+SBE3成功整合进入水稻基因组中。

[Objective]The research aimed to develop the amylose content and expand the applied scope of rice.[Method] In this experiment,transformation was conducted on Oryza sativa callus mediated by Agrobacterium and the RNAi vector constructed gene was integrated into Oryza sativa genome.[Results] Transformation of RNAi plasmid including the Oryza sativa L branching enzymes（SBE1 and SBE3）was conducted on Oryza sativa callus of Zhonghua11 mediated by Agrobacterium.After co-cultivation,screening, predifferentiation,differentiation and radication,49 transgenic plants were acquired.The Hygromycin gene fragment was amplified from the genome DNA of transgenic Oryza sativa L by PCR.[Conclusion] The prelimmary success of Oryza sativa transgene is confirmed.