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目的 构建 HAb1 8G反义 RNA表达质粒载体 .方法 用 DNA重组技术将人 HAb1 8G基因反向克隆到真核表达质粒 PCI- neo中 ,构建成 HAb1 8G反义 RNA表达质粒PCI- as HAb1 8G.用阳离子脂质体介导转染人肝癌细胞系HHCC,经 G41 8筛选后获得的克隆进行鉴定 .结果 HAb1 8G反义 RNA表达质粒载体 PCI- as HAb1 8G经限制性酶切及部分序列分析证明基因插入正确 .流式细胞仪及免疫组化 SP法染色均证实 :转染细胞 HHCC/as HAb1 8G中HAb1 8G表达为阴性 ,而转染空载体的 HHCC/neu及 HHCC均为强阳性 .结论 成功构建了 HAb1 8G反义 RNA表达质粒载体 PCI- as HAb1 8G.为进一步研究 HAb1 8G蛋白分子的生物学功能及肝癌的基因治疗研究奠定了基础
Objective To construct HAb1 8G antisense RNA expression plasmid vector.Methods Human HAb1 8G gene was reverse cloned into eukaryotic expression plasmid PCI-neo by DNA recombination technology and constructed into HAb1 8G antisense RNA expression plasmid PCI-as HAb1 8G. Cationic liposomes were transfected into human hepatocellular carcinoma cell line HHCC and identified by G41 8.Results The restriction fragment and partial sequence analysis of the HAb1 8G antisense RNA expression plasmid vector PCI-as HAb1 8G proved that the gene Insertion correct.Flow cytometry and immunohistochemical SP staining confirmed that the transfected cells HHCC / as HAb1 8G HAb1 8G expression was negative, while the transfected empty vector HHCC / neu and HHCC were strongly positive.Conclusion The successful The HAb1 8G antisense RNA expression plasmid vector PCI-as HAb1 8G was constructed, which laid the foundation for the further study on the biological function of HAb1 8G protein gene and the gene therapy research of HCC