丙型肝炎病毒NS 3不同末端在COS、NIH 3T3细胞中的转录与表达

Transfection and expression of the gene from HCV-NS 3 different terminals in COS and NIH 3 T 3 cells

目的:建立丙型肝炎病毒NS3区不同末端的COS和NIH3T3的细胞系表达系统,以探讨其慢性化及肝 癌的发病机理。方法:用脂质体共转录法,将含有丙型肝炎病毒NS3区不同末端的重组质粒pSG5neo转录至 COS和NIH3T3细胞系中表达,并用聚合酶链反应(polymerasechainreaction,PCR)和Southernblot法在DNA 水平检测。用Westernblot法在蛋白水平检测。结果:在转录的COS和NIH3T3的细胞系中,存在有丙型肝炎 病毒NS3在3'和5'基因片段,并可表达相应的抗原。结论:此表达细胞系的建立,有利于对丙型肝炎病毒的复 制,丙型肝炎慢性化及肝癌发病的研究。

Objective:Hepatitis C Virus(HCV) is the major etiological agent of posttransfusion as well as community acquired non A non B. Chronic hepatitis and persistent infections by HCV often lead to liver cirrhosis and hepatoceller carcinoma (HCC). However, HCV NS 3 region is a multifunctional and connected with DNA replication. Therefore, it is useful to research chronic Hepatitis C and HCC by constructing recombinantion Plasmid pSG 5 neo eukaryotic expression vectors of HCV NS 3 different ends and transfecting them into COS cell line and NIH 3 T 3 cell line.Methods:The different terminals of HCV NS 3 gene has been transfected into COS cells and NIH 3 T 3 cells by lipofectamine. The results of transfection were confirmed by PCR and Southern blot analysis, and the levels of their proteins in COS and NIH 3 T 3 cells were detected by Western blot analysis.Results:The cDNA fragments and proteins of HCV NS 3 different terminals in COS cell and NIH 3 T 3 cells were foond. Conclusion:The established COS and NIH 3 T 3 cell lines containing the different terminals of HCV NS 3 could be used to investigate HCV replication and the mechanism of persistent hepatitis C and HCC.