【摘要】 将细粒棘球蚴(Echinococcus granulosus,Eg.)延伸因子(Elongation factor 1,EF-1)与pGEX-6P-1构建成重组质粒并进行表达、纯化及对其特性进行鉴定。将构建好的EF-1/pGEM-T经双酶切后获取目的基因片段,定向重组于表达载体pGEX-6P-1并转化大肠杆菌(Escherichia coli)E.coliBL21,IPTG诱导表达,用蛋白亲和层析柱GSTrap FF对表达产物进行纯化,特异性蛋白解离酶PreScissionTMProtease酶切融合蛋白从中获取重组Eg.EF-1;SDS-PAGE和Western blot方法鉴定表达产物。获得的Eg.EF-1/pGEX-6P-1/E.coliBL21菌株,诱导表达融合蛋白和纯化分离得到的31 ku Eg.EF-1均能被细粒棘球蚴天然抗原免疫的兔多克隆抗血清识别。证明成功构建出具有有效表达的Eg.EF-1/pGEX-6P-1菌株,初步证实重组蛋白具有较好的抗原性。更多还原

【Abstract】 To obtain the recombinant protein Eg.EF-1 by gene engineering,the gene fragment of（Eg.EF-1） was obtained by restrictedly digesting the Eg.EF-1/pGEM-T,and subclonding into expression vector pGEX-6P-1.The Eg.EF-1/pGEX-6P-1 was constructed and the fusion protein Eg.EF-1/GST（expressed） in E.coli BL21 induced by IPTG.The Eg.EF-1/GST was purified by chromatography using GSTrap FF and the isolated 31ku Eg.EF-1 has been obtained after the fusion protein digested by the PreScission^TMProtease.Using Western blotting the Eg.EF-1 and Eg.EF-1/GST can be identified by the sera of rabbits immunized by hydatid cyst fluid from CE patients.The recombinant fusion protein（Eg.EF-1/GST） has a same epitope as the nature antigen.更多还原