pBIN438-CMV △Rep表达载体的构建及其在根癌农杆菌中的转化  

PBIN438-CMV△Rep Expression Vector and its Transformation in Agrobacterium Tumefaciens

作　　者：雷霄飞[1] 杨学领[1] 

机构地区：[1]武汉生物工程学院科技处,湖北武汉430415

出　　处：《安徽农学通报》2015年第8期13-14,共2页Anhui Agricultural Science Bulletin

基　　金：湖北省教育厅资助项目(B20129002);武汉市教育局科研资助项目(2010087)

摘　　要：用PCR方法扩增黄瓜花叶病毒部分复制酶基因(CMV△Rep),连接到PUCm-T载体上构建成克隆载体PUCm-T-CMV△Rep。用Bam HⅠ和SalⅠ分别对克隆载体PUCm-T-CMV△Rep和植物表达载体p BIN438进行双酶切,获得目的片段和线性质粒。在T4 DNA连接酶的作用下进行定向连接,构建成植物表达载体p BIN438-CMV△Rep。采用Ca Cl2冻融法将重组子导入根癌农杆菌LBA4404。经PCR和双酶切鉴定,表明重组质粒p BIN438-CMV△Rep已成功导入根癌农杆菌中。Cucumber mosaic virus partial replicase gene were amplified by PCR（CMV Rep）,connected to the PUCm-T vector to construct the cloning vector of PUCm-T-CMV Rep. With BamH I and Sal I of PUCm-T-CMV cloning vector Rep and the plant expression vector pBIN438 were digested,obtained fragment and linear plas？mid. Directional connection in T4 DNA ligase,a plant expression vector was constructed by pBIN438-CMV Rep. Using CaCl2 freeze-thaw method the recombinant plasmid into Agrobacterium LBA4404. By PCR and double enzyme diges？tion showed that the recombinant plasmid,pBIN438-CMV Rep has been successfully introduced into Agrobacteri？um tumefaciens.

关 键 词：复制酶基因 载体构建 农杆菌 黄瓜花叶病毒 

分 类 号：S188[农业科学—农业基础科学]