编码霍乱肠毒素B亚单位基因植物表达载体的构建  

Construction of plant expression vectors containing the gene encoding cholera toxin B subunit

作　　者：彭志强[1] 俞守义[1] 余迪求[2] 贺竹梅[2] 

机构地区：[1]第一军医大学流行病学教研室,广东广州510515 [2]中山大学生物工程研究中心,广东广州510275

出　　处：《第一军医大学学报》2002年第8期736-738,共3页Journal of First Military Medical University

摘　　要：目的构建含霍乱肠毒素B亚单位(CTB)基因的植物双元表达载体。方法采用高保真PCR方法调出CTB基因,定向克隆到中间载体pRTL2,经测序证实核酸序列正确后,再亚克隆到含植物双元表达载体pBI121上;采用电击法,将含CTB的植物表达载体转入根癌农杆菌中,并进行酶切证实。结果PCR扩增的CTB片断亚克隆到中间载体,得到pRCTB和pRCTBK,测序证实CTB核酸序列正确;再与根癌农杆菌双元载体pBI121连接,获得含CTB基因的植物双元表达载体pBI-CTB和pBI-CTBK,转入根癌农杆菌的载体,酶切证实正确。结论采用正确技术路线,构建了含CTB基因的植物表达载体,为下一步的转基因植物表达研究打下了坚实的基础。Objective To construct the plant expression vector containing the nucleotide sequence encoding cholera toxin B (CTB) subunits. Method Using high-fidelity PCR, we amplified CTB genes that were then subcloned into the transition vec-tor pRTL2. Following confirmation of the CTB nucleotide sequence, the vector was subcloned into the plant vector pBI121 that was subsequently transferred into Agrobacterium tumefaciens LBA4404 by electroporation. Results CTB DNA that was ligated into the transition vectors resulted in the 2 vectors designated as pRCTB and pRCTBK. After the 2 vectors were ligated into the plant binary vector pBI121 respectively, new plant binary vectors, namely pBI-CTB and pBI-CTBK, were produced. Analysis with restriction endonucleases confirmed successful transfer of pBI-CTB and pBI-CTBK into Agrobacterium tumefa-ciens LBA4404. Conclusion With appropriate technological strategy, the plant binary expression vectors encoding CTB have been constructed, which facilitates further investigation of CTB protein expressions in transgenic plant.

关 键 词：霍乱肠毒素 亚克隆 多聚酶链式反应 植物表达载体 

分 类 号：R346[医药卫生—基础医学]