幽门螺杆菌全长cagA基因和霍乱毒素B亚单位基因的克隆与表达

Cloning and expression of the CagA gene of Helicobacter pylori and the gene of cholera toxin subunit B

目的 构建表达幽门螺杆菌 (Hp)细胞毒素相关蛋白 (CagA)及粘膜免疫佐剂霍乱毒素B亚单位 (CTB)的重组质粒 ,并在大肠杆菌中表达获得基因重组蛋白。方法 用PCR方法从幽门螺杆菌扩增CagA基因片段 ,从霍乱弧菌扩增CTB基因片段 ,将它们转入原核载体质粒pGEMEX 1,在大肠杆菌DH5α中克隆 ,并在JM10 9DE3中表达。结果 重组质粒pGEMEX CTB的全长序列经分析与Gen Bank公布的序列相符 ;各表达蛋白经SDS PAGE分析 ,相对分子量与文献相符 ;重组蛋白经Westernblot检测有较强的抗原性。

Objective To construct the recombinant plasmid of Helicobacter pylori(Hp) CagA and mucasal adjuvant cholera toxin subunit B (CTB) and express fusion proteins in E.coli JM109DE3. Methods Gene encoding CagA was amplified from Hp chromosomal DNA and CTB from Vibrio cholerae. The genes were inserted into the prokaryotic expression vector pGEMEX-1. Recombinant CagA and CTB were expressed in E.coli JM109DE3. Results The DNA sequence of recombinant pGEMEX-CTB was identical with that published by GenBank publicized. The relative molecular mass (M r) of fusion protein was corresponding to the M r shown in SDS-PAGE. Western blot analysis of the recombinant CagA confirmed its strong immunogenicity. Conclusion The recombinant protein may function as a potential vaccine or as a diagnostic antigen.