幽门螺杆菌重组Bb-hpaA-vacA疫苗的构建

Construction of recombinant Bb-hpaA-vacA vaccine of Helicobacter pylori

目的构建幽门螺杆菌(Hp)重组双歧杆菌(Bb)Bb-hpaA-vacA疫苗。方法通过PCR分别扩增hpaA和va-cA抗原编码基因,然后采用基因拼接法(gene SOEing)剪接hpaA和vacA,得到hpaA-vacA融合基因;将该融合基因定向克隆到大肠埃希菌-双歧杆菌穿梭表达载体pGEX-1λT,构建重组质粒pGEX-hpaA-vacA,电穿孔法将该质粒导入Bb,构建幽门螺杆菌重组hpaA-vacA疫苗,然后用SDS-PAGE和Western blotting鉴定表达的重组蛋白。结果 PCR成功扩增出分子量约为1 500bp的hpaA-vacA融合基因,双酶切证实hpaA-vacA融合基因成功插入pGEX-1λT中,并成功转化入双歧杆菌,而且重组蛋白能在双岐杆菌中得到正确表达,Western blotting显示重组蛋白具有免疫原性。结论成功构建螺杆菌rBb-hpaA-vacA疫苗,为该疫苗的进一步研究奠定了基础。

To construct the recombinant BbhpaAvacA vaccine of Helicobacter pylori, hpaA and vacA antigen genes were amplified by PCR and the fusion gene hpaAvaeA was obtained with gene SOEing. Then the fusion gene was cloned into Escherichia coliBifidobacteria shuttle plasmid pGEX-lkT to construct pGEXhpaAvacA. The recombinant plasmid was electroporated into Bifidobacteria bifidum （Bb） to construct rBb hpaAvacA vaccine. The recombinant protein was analyzed by SDSPAGE. Its immunogenicity was identified by Western blotting. A 1500bp fusion gene of hpaA-vacA was successfully amplified by PCR and cloned into pGEX,T by restriction analysis, and the rBb hpaA vacA vacci,ze was successfully constructed by PCR and restriction analysis. The recombinant protein could be expressed in B. bifidium and it has immunogenicity. In this way, the rBb hpaAvacA vaccine of Helicobacter pylori is successfully constructed, which lays the experimental foundation of exploitation and utilization of this vaccine.