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目的 构建HCV包膜糖蛋白E2与His标签融合的真核表达载体并在CHO细胞中表达 ,为研究HCV包膜糖蛋白的功能奠定基础。方法 利用PCR技术从HCV基因组序列中扩增出编码HCV包膜糖蛋白E2的基因 ,将其插入原核表达载体pET2 8(a)载体中 ,构建pET2 8(a) HCVE2 ,从pET2 8(a) HCVE2上获得His HCVE2融合基因 ,插入pcDNA3 1,构建表达HCV包膜糖蛋白E2与His标签融合蛋白的真核表达载体pcDNA3 1 His E2。用脂质体介导法将pcDNA3 1 His E2转染CHO细胞 ,通过间接免疫荧光、Westernblot检测融合蛋白在CHO细胞内的表达。结果 转染HCVE2与His融合基因的真核表达载体的CHO细胞内有融合蛋白的表达。结论 与His标签融合的HCV包膜糖蛋白E2能够在CHO细胞内表达。
Objective To construct an eukaryotic expression vector fused HCV envelope glycoprotein E2 and His tag and lay the foundation in CHO cells for the study of the function of HCV envelope glycoprotein. Methods The HCV envelope glycoprotein E2 gene was amplified from HCV genome sequence by PCR and inserted into the prokaryotic expression vector pET28 (a) to construct pET28 (a) HCVE2. HCVE2 fusion protein His HCVE2 inserted into pcDNA3 1, the expression of HCV envelope glycoprotein E2 and His tag fusion protein eukaryotic expression vector pcDNA3 1 His E2. The CHO cells were transfected with pcDNA3 1 His E2 by lipofectamine. The expression of the fusion protein in CHO cells was detected by indirect immunofluorescence and Western blot. Results The fusion protein was expressed in CHO cells transfected with eukaryotic expression vector of HCVE2 and His. Conclusion The HCV envelope glycoprotein E2 fused to His-tag was expressed in CHO cells.