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| 摘要: |
| 利用DNA重组技术将丙型肝炎病毒（HCV）H77株E1E2囊膜蛋白基因插入逆转录病毒载体pBABE-puro中，构建成重组逆转录病毒载体pBABE-puro-E1E2，用重组逆转录病毒载体与pVSVg质粒磷酸钙共沉淀法转染293T细胞,包装逆转录病毒假病毒。用包装的假病毒感染SP2/0细胞，经嘌呤霉素筛选阳性细胞后进行流式细胞仪技术（FACS）分析，结果表明,HCV E1E2基因在SP2/0细胞膜上成功表达。将表达E1E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠，经FACS分析免疫鼠血清，成功诱导小鼠产生了抗HCV E1E2蛋白的抗体，Western blot检测结果表明该抗体能与原核系统表达的E2蛋白结合。 |
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| 基金项目:国家自然科学基金资助项目（30340027） |
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| Expression of hepatitis C virus E1E2 envelope protein gene in eukaryotic cells and animal immune experiment |
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| Abstract: |
| The recombinant retroviral vector pBABE-puro-E1E2 was constructed by inserting full-length HCV E1E2 gene of H77 strain into pBABE-puro.Both the recombinant retroviral vector and pVSVg plasmid were transfected into eukaryotic cells 293T by calcium phosphate transfection method.And then,the pseudovirus were produced.The pseudovirus infected eukaryotic cells SP2/0 and E1E2 protein was expressed.E1E2 protein was detected by puromycin-resistant and FACS analysis.BALB/c mice were injected in abdomen with SP2/0 cells expressing E1E2 protein.Anti-HCV E1E2 antibody was screened by FACS.Moreover,the antibody was also analyzed by Western blot using E2 protein antigen which was expressed in E.coli.The results showed that HCV E1E2 protein was expressed in SP2/0 cells' envelope successfully.FACS could detect specific anti-E1E2 antibody in SP2/0 cells immune mouse serum.Western blot analysis showed that SP2/0 cells immune mouse serum could react specially to E2 protein expressed by E.coli. |
| Key words: hepatitis C virus envelope protein gene retroviral vector pseudovirus FACS |
