中文摘要：

目的构建丙型肝炎病毒（HCV）全基因重组质粒，并建立其转染细胞系。方法双酶切质粒JFH1，回收HCV DNA，分别与真核表达载体pIRESneo和逆转录病毒载体pLNCX连接，构建重组质粒pIRESneo—HCV和pLNCX—HCV。将重组质粒pIRESneo—HCV转染BHK细胞，pLNCX—HCV转染pA317细胞，通过PCR、间接ELISA和免疫荧光试验鉴定转染细胞。结果重组质粒PCR和酶切鉴定证明构建正确。转染细胞上清经PCR检测均可见目的基因条带；ELISA结果表明，转染细胞冻融上清均可与抗HCV小鼠血清发生强的结合反应；转染细胞均可与抗HCV小鼠血清反应，产生荧光，但荧光不多且斑点不亮。结论已成功构建HCV全基因重组质粒，并建立了可表达HCV蛋白的转染细胞系，为转基因动物模型的建立奠定了基础。

英文摘要：

Objective To construct a recombinant plasmid with full-length hepatitis C virus （HCV） gene and establish a transfected cell line. Methods Plasmid JFH1 was digested with Age 1 and Xba 1, and HCV DNA was recovered and inserted into plasmids pIRESneo and pLNCX respectively. The constructed recombinant plasmids pIRESneo-HCV and pLNCX-HCV were transfected to BHK and pA317 cells respectively, and the transfected cells were identified by PCR, indirect ELISA and IFA. Results PCR and restriction analysis proved that recombinant plasmids plRESneo-HCV and pLNCX-HCV were constructed correctly. The target gene bands were detected by PCR in the culture supematants of transfected BHK and pA317 cells. ELISA proved strong binding activities of superuatants of transfected BHK and pA317 cells after freeze-thawing with murine antisera against HCV. IFA showed specific reactions of transfected BHK and pA317 cells with murine antisera against HCV, however, the fluorescent foci were in a small number, and their brightness was low. Conclusion The recombinant plasmid with full-length HCV gene was successfully constructed, and the transfected cell lines were established, which laid a foundation of establishing transgenic animal model.