幽门螺杆菌空泡毒素表达系统的构建及其鉴定

采用常规酚 氯仿法提取幽门螺杆菌 (Helicobacterpylori,Hp)临床菌株Y0 6株基因组DNA ,用PCR扩增空泡毒素 (vacA)基因 ,T A克隆后测序 ,亚克隆构建表达载体 ,采用Ni NTA亲和层析法提纯表达产物 ,SDS PAGE和Westernblot分别检测表达产物的分子量和免疫性。所构建的表达系统能表达vacA ,其产物可与相应抗体结合。重组vacA能有效地诱导家兔产生抗体 ,该抗体也能与重组vacA发生结合反应。

Construction of the Expression System of Helicobacter pylori Vacuolating Cytotoxin Gene and Identification of Immunity of the Recombinant Protein

The genomic DNA of a clinical strain Y06 of Helicobacter pylori was prepared by routine phenol-chloroform method. The whole vacA gene fragment was amplified by PCR and then sequenced after T-A cloning. The expression vector of vacA gene was constructed through sub-cloning. The expression product was extracted and purified by Ni-NTA affinity chromatography. The molecular weight and immunity of recombinant vacA were determined by SDS-PAGE and Western blot, respectively. The constructed expression system is able to produce recombinant vacA and the product could combine with the corresponding antibody. The rvacA could efficiently induce rabbit to produce specific antibody and the antibody was also able to combine with the rvacA. ;