【摘要】 目的　构建含幽门螺杆菌 (Hp) ure A、ure B基因重组质粒 ,测定、分析其核酸序列及推定的氨基酸序列 ,在 E.coli中高效表达两基因 ,为检测试剂和疫苗研究提供基础依据和抗原。方法　PCR法扩增 Hp菌株 HPSH4的 ure A、ure B基因 ,分别与 p GEX- 2 T载体连接并转化大肠杆菌JM10 5 ,测定克隆基因的核酸序列并与 Gen Bank公布的国外 5株 Hp菌株 (2 6 6 95 ,HPK 5 ,J99,CPM6 30 ,M6 0 398)的 ure A、ure B基因作序列比较 ,以 IPTG诱导 ,ure A、ure B基因在 E.coli中高效表达。结果　 ure A核酸同源性 96 .3%～ 98.1% ,氨基酸同源性 98.3%～ 10 0 % ;ure B核酸同源性95 .8%～ 98.0 % ,氨基酸同源性 96 .4 %～ 99.6 %。以 IPTG诱导 ,在 E.coli中表达 rure A融合蛋白5 5 6 0 0 D,rure B融合蛋白 85 5 0 0 D。结论　不同地区 Hp菌株的 ure A、ure B存在程度不同的变异 ,克隆并表达 HPSH4的 ure A、ure B与 Gen Bank已公布的多数 Hp菌株有极高同源性 ,在 E.coli以融合蛋白高效表达 rure A和 rure B更多还原

【Abstract】 Objective To obtain rureA, rureB of H.pylori for developing vaccines and applying in diagnostic kits by cloning, sequencing and expression of ureA, ureB genes from H.pylori in E.coli. Methods The ureA, ureB genes of H.pylori (HPSH4) were amplified by PCR. The PCR products were cloned into plasmid pGEX 2T respectively. The cloned ureA, ureB genes were sequenced and compared with GenBank strains (26695,HPK5, J99,CPM630,M60398). The cloned genes were expressed in E.coli by induction with IPTG. Results Sequencing analysis showed that 96.3%～98.1% of nucleotides, 98.3%～100% of putative amino acid and 95.8%～98.0% of nucleotides, 96.4%～99.6% of putative amino acid are homologous among cloned ureA,ureB and 5 Hp strains in GenBank. In E.coli, the pGEX 2T/ureA and pGEX 2T/ureB expressed rureA 55600D, rureB 85500D in fusion protein, respectively. Conclusions The cloned ureA, ureB gene expressed in E.coli are highly homologous with most of the GenBank Hp strains. The basic data and antigen for diagnostic kits and vaccine were obtained.更多还原