Construction of survivin gene vector and its expression in human hepatocellular carcinoma cell line HepG2

    CHENG Shengª²Quan, WANG Wenª²Liang, YAN Wei

    1Department of Pathology, Xijing Hospital, 2Department of Pathology, School of Basic Medicine, Fourth Military Medical University, Xi¡¯an 710033, China

    ¡¾Abstract¡¿ AIM£º To clone coding sequence of survivin (SVV) gene and to construct its eukaryotic expression vector for exploring the roles of SVV gene in the carcinogenesis of hepatocellular carcinoma. METHODS: SVV gene was amplified from HLª²60 by RTª²PCR and the fragments of the cDNA were cloned into eukaryotic expression vector pcDNA3.0 by ligating the fragments into BamHI and XhoI site. The recombinant plasmid pcDNA3.0ª²SVV was identified by DNA sequence and restriction analysis. The gene transfection mediated by the lipofectin was used to introduce the eukaryotic expression vector of pcDNA3.0ª²SVV into human hepatocellular carcinoma cell line HepG2. After selected with G418, resistant colonies were obtained. The immunohistochemical staining and RTª²PCR was analyzed. RESULTS: A 445 bp DNA fragment was amplified with RTª²PCR. Sequence and restriction analysis showed that the recombinant plasmid pcDNA3.0ª²SVV was constructed successfully. CONCLUSION: Human SVV gene has been successfully cloned. The sequence analysis reveals that it is 100% homologous to data published in GenBank. The construction of the recombinant plasmid pcDNA3.0ª²SVV will be helpful in the further study of this SVV gene in the development of hepatocellular carcinoma. ......