目的构建ＨＣＶ５ＮＣＲ片段调控荧光素酶表达质粒，并在ＨｅｐＧ２细胞中表达。方法通过ＰＣＲ扩增，获得中国人ＨＣＶ基因组５非编码区（ｎｏｎｃｏｄｉｎｇｒｅｇｉｏｎ，ＮＣＲ）完整序列与Ｃ区部分序列的目的基因片段（５ＮＣＲ－Ｃ片段）。将此片段插入ｐＧＬ３荧光素酶报告载体的荧光素酶基因起始密码上游，构建受５ＮＣＲ片段调控的荧光素酶表达质粒。应用脂质体介导基因转染技术将８个质粒转染ＨｅｐＧ２肝癌细胞。结果经鉴定获得８个均为正向插入的阳性克隆。序列分析结果表明插入片段与中国人ＨＣＶ（Ⅱ型）序列基本一致，其荧光素酶基因起始密码子已去除且未改变编码框。有一个质粒能够表达荧光素酶活性，其绝对值达１１４１．９±１５１．１ｍＶ，是ｐＧＬ３报告载体荧光素酶活性的２０％。结论当质粒１μｇ、Ｌｉｐｏｆｅｃｔｉｎ６μｌ，Ｌｉｐｏｆｅｃｔｉｎ－质粒复合物与细胞孵育４小时时可获得最佳转染效果 Objective To construct a luciferase expression plasmid for HCV 5 ’NCR fragment and express it in HepG2 cells. Methods The target gene fragment (5NCR-C fragment) of the 5 ’noncoding region (NCR) complete sequence and the C region partial region of Chinese HCV genome was obtained by PCR amplification. This fragment was inserted upstream of the luciferase gene start codon of the pGL3 luciferase reporter vector to construct a luciferase expression plasmid regulated by a 5? NCR fragment. Eight plasmids were transfected into HepG2 hepatoma cells by liposome-mediated gene transfection. Results Eight positive positive clones were identified. The results of sequence analysis showed that the inserted fragment was basically identical with the Chinese HCV (type Ⅱ) sequence, and the luciferase gene start codon was removed without changing the coding sequence. One plasmid expressed luciferase activity with an absolute value of 1141.9 ± 151.1 mV, which is 20% of the luciferase activity of the pGL3 reporter vector. Conclusion When the plasmid 1μg, Lipofectin 6μl, Lipofectin-plasmid complex incubated with cells for 4 hours when the best transfection results can be obtained