非结构蛋白_____HCV；_基因；_非结构蛋白；

HCV core真核表达质粒pcDNA3.1(-)/core的构建

http://www.100md.com 《广东药学院学报》 2005年第1期

非结构蛋白,,HCV；,基因；,非结构蛋白；,载体,HCVcore真核表达质粒pcDNA3.1(-),core的构建,1材料与方法,2结果,3讨论,参考文献

     摘要目的构建HCV core基因真核表达质粒，为进一步研究和解析HCV诱发人不死化肝细胞癌化的机制,HCV 感染的检测及疫苗和药物研发做好前期基础。方法将含HCV全长基因的pBRTM/HCV1-3011质粒于大肠杆菌JM109内扩增；提取pBRTM/HCV1-3011质粒;从pBRTM/HCV1-3011质粒中PCR扩增出HCV core片段并将其插入pGEM-T克隆载体；再与表达载体pcDNA3.1(-)重组,以得到重组的真核表达载体pcDNA3.1(-)/core；最后限制性酶切鉴定HCV core表达载体。结果从pBRTM/HCV1-3011质粒中扩增出的HCV core片段大小正确，经测序证明其碱基序列为编码目的基因的正确序列；凝胶电泳结果证明已将此片段克隆到pcDNA3.1/core内。结论成功的构建了HCV core基因的真核表达载体pcDNA3.1(-)/core。

    关键词 HCV；基因；非结构蛋白；载体

    Construction of eukaryotic plasmid pcDNA3.1(-)/core containing the core gene of HCV

    LIN Junwu,JIANG Jing, WANG Zhipeng, XU Xiaoliang, LIN Shaoqiang, HUANG Zeqi, WEI Jing, LI Xiaolan

    (Department of Microbiology and Immunology, Medical College, Jinan University, Guangzhou 510632, China)

    Abstract Objective To construct the eukaryotic plasmid containing the core gene of HCV, preparing for the research in the mechanism of the cancerization induced by HCV. Methods After expanding plasmid pBRTM/HCV1-3011 (containing the whole genome of HCV) through coliform JM109, we PCR the core gene with pBRTM/HCV1-3011 as the template; Insert the core segment into the clone plasmid pGEM T. Digest plasmid pcDNA3.1(-)and pGEM T/core with EcoRI and BamHI, then recycle the digested pcDNA and core segment. The pcDNA and core segment were circularized with T4 DNA ligase. Results The PCR results of HCV core has correct size and sequences. Conclusion The eukaryotic plasmid pcDNA3.1(-)/core has successfully been constructed. ......

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