丙型肝炎病毒NS_5B在Hub-7细胞中的转染与表达

Transfection and expression of HCV-NS_5B gene in Huh-7 cells

目的建立丙型肝炎病毒（ＨＣＶ）ＮＳ５Ｂ表达的人肝细胞系。方法以脂质体共转染ＮＳ５ＳＢ基因人Ｈｕｂ－７细胞，以ＰＣＲ和Ｓｏｕｔｈｅｒｎ ｂｌｏｔ在ＤＮＡ水平检测转染的结果，以ｗｅｓｔｅｒｎ ｂｌｏｔ证实了Ｈｕｂ－７细胞中有ＮＳｕＢ的蛋白表达。结累在转染过ＮＳＳＢ质粒的细胞系中有ＮＳＳＢ基因的存在和ＨＣＶ－ＲＮＡ多聚酶的表达。结论我们在Ｈｕｂ－７细胞中建立了ＨＣＶ－ＲＮＡ多聚酶表达系统，它有助于研究ＨＣＶ复制机制和为基因治疗丙型肝炎打下基础。

Objective Hepatitis C virus(HCV) is the major cause of blood-borne non-A, non-B hepatitis. An RNA-dependent RNA polymerase is encoded by HCV non-strUctural protein SB(NSSB) gene. A full-length HCV RNA has been transfected into human hepatoma cells, and HCV NS5B has been expressed in E.Coli purified. But human hepatocyte line with high expression of NSSB has not been established yet. Methods The NSSB gene has been transfected into Hub-7 cells by Lipofectamine. The results of transfection were confirmed by PCR and Southern blot analysis, and the level of the NSsB protein in Hub-7 cells was detected by Western blot analysis. Results There were the NSSB sequence and the expression of HCV-RNA polymerase in Hub-7 cells transfected NSSB plasmids. Conclusions We have established a HCV RNA polymerase expression system in Hub-7 cells that can be used to study the mechanism of HCV replication and to test gene therapy of hepatitis C.