豇豆胰蛋白酶抑制剂基因(CpTI)的克隆及其植物表达载体的构建  

Cloning of Cowpea Trypsin Proteinase Inhabitor Gene and Construction of Its Plant Expression Vector

作　　者：罗娟[1] 王秀敏[2] 韩烈保[2] 刘君[2] 曾会明[2] 吴云锋[1] 

机构地区：[1]西北农林科技大学动物科技学院,陕西杨凌712100 [2]北京林业大学草坪研究所,北京100083

出　　处：《西北农业学报》2008年第3期16-19,共4页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：教育部“新世纪优秀人才”支持计划

摘　　要：用CTAB法提取豇豆叶片总DNA后,根据基因两端的保守序列设计引物,经PCR方法扩增得到CpTI基因,将其克隆到pMD-18T载体上,酶切并测序鉴定,将该基因登陆到GeneBank(NO.EU088405)并进行同源性鉴定,证明该基因为丝氨酸蛋白酶抑制剂基因家族中的一员。在此基础上,将CpTI基因用限制性内切酶BamHⅠ/SacⅠ切下后,克隆到pCUbi1303的UBI启动子和NOS终止子之间。经PCR和酶切鉴定,得到了该基因的真核表达载体pCUbiCpTI1303,为下一步的转基因做好了准备。Genomic DNA in the leaves of the cowpea was extracted with CTAB.The CpTI gene was amplified by PCR with the conserved sequence acted as primers,then cloned to pMD-18T vector and detected the recombinant by restriction enzyme method.The sequence has been registered in GeneBank（NO.EU088405）.It is one of serine protease inhibitor gene family.After digesting by BamHⅠand SacⅠ,the gene was cloned to pCUbi1303 between the promoter UBI and the terminator NOS.After detected by restriction enzyme and PCR,it indicated that the plant expression vector pCUbiCpTI1303 was constructed successfully,which provides a basis for the following transgene.

关 键 词：豇豆胰蛋白酶抑制剂基因(CpTI) 克隆 植物表达载体 

分 类 号：Q943.2[生物学—植物学]