EV71病毒3C蛋白真核表达载体的构建及表达

Construction of pcDNA3.0-Flag-3C Plasmid and Its Expression of EV71 3C Protein in HeLa Cells

[目的]构建EV71病毒3C蛋白的真核表达载体,并验证其在细胞中的正确表达。[方法]采用RT-PCR技术,从EV71病毒基因组RNA中扩增3C基因,克隆到真核表达载体pc DNA3.0-Flag上,并转染He La细胞,通过Western blot和免疫荧光验证3C蛋白的表达。[结果]酶切及测序鉴定显示,EV71病毒3C真核表达载体构建成功。Western blot和免疫荧光结果证实3C蛋白在He La细胞中成功表达。[结论]该研究成功构建了真核表达载体pc DNA3.0-Flag-3C,为进一步研究EV71病毒3C蛋白生物学功能奠定了一定基础。

[Objective]To construct the eukaryotic expression vector of EV71 3C protein and investigate its expression in HeLa cells.[Method]Fragment of 3C was amplified from total RNA of EV71 genome and cloned into eukaryotic expression vector pcDNA3.0-Flag.After the recombination vector was transfected into HeLa cells,Western blot and indirect immunofluorescent assay were performed to confirm the 3C protein expression.[Result]Enzyme digestion and sequencing analysis showed that eukaryotic expression vector pcDNA3.0-Flag-3C was constructed correctly.The results of Western blot and indirect immunofluorescent indicated 3C protein was expressed in HeLa cells.[Conclusion]This research successfully constructed eukaryotic expression vector pcDNA3.0-Flag-3C,which might provide the foundation of research about the biological function of EV71 3C protein.