HCV非结构蛋白3反式激活基因6转染肝癌细胞的基因表达谱芯片分析.docxVIP

HCV非结构蛋白3反式激活基因6转染肝癌细胞的基因表达谱芯片分析 作者：邵清，成军，王琳，张健，卢成哲 【关键词】 丙型肝炎病毒  Screening of genes differentially expressed in HepGcells transfected with HCV NS3TPby microarray assay  【Abstract】 AIM: To study the difference in gene expression in human hepatoblastoma cell line HepGcells transfected with HCV NS3TP6expressing plasmid and to further elucidate its molecular biological mechanism. METHODS: Sequence specific primers were designed and synthesized and the NS3TP6coding DNA fragment was amplified with polymerase chain reaction (PCR) technique using pBRTM3011 plasmid containing the full length of HCVH cDNA as the template. The expression vector of (-)NS3TPwas constructed by routine molecular biological methods. cDNA microarray technology was employed to detect the mRNA from the HepGcells transfected respectively with (-)NS3TPand (-) using lipofectamine. RESULTS: The expression vector was constructed and confirmed by restriction enzyme digestion and DNA sequencing analysis. The expression of NS3TPprotEin was confirmed by Western blot hybridization with single chain variable region (scFv) antibody. High quality mRNA and cDNA were prepared and successful microarray screening was conducted. The scanning results indicated that among the 115genes which were obtained from gene expression profile analysis,were different, of which genes were upregulated andgenes were downregulated in NS3TP6expressing HepGcells. CONCLUSION: cDNA microarray technology is successfully used to screen the genes differentially expressed in NS3TP6expressing HepGcells, which brings some new clues for the study of the molecular mechanism involved in the HCV infection and hepatocellular carcinoma development induced by HCV NS3TPprotEIn.  【Keywords】 hepatitis C virus；NSprotein；cDNA microarray  【摘要】 目的: 为了 研究 NS3TP6可能的分子生物学功能，我们 应用 基因芯片技术，检测丙型肝炎病毒(HCV)非结构蛋白(NS3)反式激活基因NS3TP6的表达对肝母细胞瘤细胞HepG2基因表达谱的 影响 . 方法 ：以分子生物学技术构建NS3TP6的真核表达载体(-)NS3TP6，以表达质粒(-)NS3TP6转染HepG2细胞，以空载体(-)为平行对照，制备转