【作者】 曲云鹏； 刘建国； 柴巧学； 杨德琴； 管晓燕； 韩琪； 张剑； 白国辉；

【Author】 QU Yun-peng1,2,LIU Jian-guo1,CHAI Qiao-xue1,YANG De-qin1,GUAN Qian-yan1, Han qi1,ZHANG Jian1,BAI Guo-hui1.(1.Dept.of Oral medcine,The Affiliated Stomatology Hospital of Zunyi Medicine,2.The Affiliated Stomatology Hospital,Wenzhou Medical College,Wenzhou 325027,China)

【机构】 遵义医学院附属口腔医院口腔内科； 温州医学院附属口腔医院儿童牙科；

【摘要】 目的:将变异链球菌表面蛋白PAcA区编码基因spap/A和葡糖基转移酶催化区(CAT)编码基因gtfB/CAT的序列克隆到真核载体pVAX1中,构建spap/A-gtfB/CAT嵌合质粒pVAX1-SG,并观察其在哺乳动物细胞COS-7中的表达情况。方法:通过基因重组技术构建真核表达质粒pVAX1-SG;通过脂质体转染法将其转染至COS-7细胞中,然后经免疫组化SABC法检测其在细胞中的表达。结果:真核表达质粒pVAX1-SG经酶切分析证实携带spap/A和gtfB/CAT片段,经测序测定,碱基对同源性达100%;pVAX1-SG转染的细胞胞质呈褐色染色,阴性对照细胞胞质无着色。结论:成功构建真核表达质粒pVAX1-SG,所携带的基因序列正确,能够在真核细胞COS-7中正确表达融合蛋白。更多还原

【Abstract】 OBJECTIVE:To evaluate the expression of recombinant plasnnid pVAX1-SG of surface protein antigen A and glucosuiteansferases CAT of Streptococcus mutans in mammalian cells COS-7.METHODS:To construct the fusion eularyotic plamid pVAX1- SG carrying encoding gene of spap/A and gtfB/CAT of Streptococcus mutans and to introduce the plasmid into COS-7 cells by lipofetamine reagent.The transient expressed protein was detected by immunochemistry technique in COS-7 cells.RESULTS:The positive expression was detected in plasma of the cells which were transfected with recombinant plamid pVAX1-SG.The cells which were transfected with pVAX1 and pVAX1- gtfB/CAT were negative.CONCLUSION:Fusion eukaryotic plasmid pVAX1- SG is successfully construted and confirmed by sequencing and enzymes digestion.The expressed protein locates in the plasma and the protein is able to combine with anti-GTF antibody.The expressed protein has the antigencity and the recombinant plasmid pVAX1- SG is a candidate vaccine.更多还原