甜菜内向整流K^+通道BvAKT1基因RNAi载体的构建  

Construction of RNAi Expression Vector of BvAKT1 Gene encoding Inward Rectifier K^+ Channel in Sugar Beet

作　　者：伍国强[1] 刘海龙[1] 李胜胜 蔺丽媛 谢玲玲 Wu Guoqiang;Liu Hailong;Li Shengsheng;Lin Liyuan;Xie Lingling(School of Life Science and Engineering,Lanzhou University of Technology,Lanzhou,730050)

机构地区：[1]兰州理工大学生命科学与工程学院,兰州730050

出　　处：《分子植物育种》2019年第1期112-118,共7页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31460101);兰州理工大学"红柳杰出人才"培养计划项目(J201404)共同资助

摘　　要：植物细胞质中维持较高的K+/Na+是植物抵御盐胁迫的有效策略之一。内向整流K+通道AKT1(Arabidopsis K+transporter)在这一过程中发挥着重要作用。本研究以嗜盐作物甜菜(Beta vulgaris L.)幼苗为材料,提取其根总RNA,采用RT-PCR方法扩增得到甜菜K+通道BvAKT1基因的RNAi靶片段,以中间载体pHANNIBAL和植物表达载体pART27为基础,采用酶切和连接的方法构建了由CaMV 35S启动子驱动、含BvAKT1基因片段反向重复序列的RNAi (RNA interference)植物表达载体pART-BvAKT1-RNAi,采用冻融法将其分别导入根癌农杆菌GV3101和发根农杆菌LBA9402,对揭示BvAKT1在甜菜K+吸收及离子稳态平衡中的作用研究具有推动作用。Maintaining higher ^K+/Na^+in plant cytoplasm is one of the effective strategies for plants to resist salt stress. The inward rectifier K+channel AKT1(Arabidopsis K+transporter) plays an important role in this process.In the present study, the seedling of halophilic crop Beta vulgaris L. was used as the material and the total RNA of its root was extracted. The RNAi target fragment of BvAKT1 gene in K^+channel of sugar beet was obtained by RT-PCR amplification. Based on the intermediate vector pHANNIBAL and the plant expression vector pART27,the RNAi(RNA interference) plant expression vector pART-BvAKT1-RNAi with inverted repeats of BvAKT1 driven by the promoter CaMV 35 S was successfully constructed by restriction enzyme digestion and ligation methods. Furthermore, the recombinant plasmid was transformed into Agrobacterium tumefaciens GV3101 and Agrobacterium rhizogenes LBA9402 by freezing and thawing method, respectively. This work could promote the research on the mechanism of BvAKT1 in K+uptake and ions homeostasis of sugar beet.

关 键 词：甜菜 内向整流K^+通道 K^+吸收 RNA干扰 

分 类 号：S566.3[农业科学—作物学]