【摘要】 目的探讨乙型肝炎病毒(HBV)前S2蛋白(preS2)对人酰基蛋白硫酯酶1(APT1)启动子的作用。方法生物信息学方法确定人APT1启动子序列。PCR扩增人APT1启动子和HBV preS2基因,分别插入pGL3和pcDNA3.1(-)质粒构建人APT1启动子荧光素酶报告基因质粒pGL3-APT1和HBV preS2真核表达质粒pcDNA3.1(-)-preS2。将pGL3-APT1和pcDNA3.1(-)-preS2共转染人肝癌细胞系HepG2,然后通过检测细胞荧光素酶活性来评价preS2对人APT1基因启动子的作用。数据用独立样本t检验分析。结果测序结果证实pcDNA3.1(-)-preS2与pGL3-APT1与实验设计相符。pGL3-APT1的荧光素酶活性是阳性对照质粒pGL3-Control的荧光素酶活性的1.2倍(P<0.01)。pcDNA3.1(-)-preS2与pGL3-APT1共转染HepG2细胞的荧光素酶活性为无preS2基因质粒pcDNA3.1(-)与pGL3-APT1共转染HepG2细胞荧光素酶活性的2.6倍(P<0.01)。结论本研究克隆的人APT1启动子序列具有高启动子活性;HBV preS2可激活人APT1启动子。更多还原

【Abstract】 Objective To investigate the trans-regulative effect of hepatitis B virus(HBV)preS2on the promoter of human acyl protein thioesterase 1(APT1)gene.Methods The promoter sequence of human APT1 gene was identified applying the software of bioinformatics.The APT1 promoter and HBV preS2 gene were amplified with PCR and cloned into pGL3 and pcDNA3.1(-)plasmids to construct the luciferase reporter gene plasmid of human APT1 gene promoter pGL3-APT1 and the preS2 eukaryotic expression plasmid pcDNA3.1(-)-preS2,respectively.The effect of the preS2 on the human APT1 gene promoter was examined by cotransfecting hepatocellular carcinoma cell HepG2 with pGL3-APT1 and pcDNA3.1(-)-preS2 and measuring luciferase activities of the HepG2 cells.The statistical data were analyzed with independent-samples t test.Results Both plasmids of pGL3-APT1 and pcDNA3.1(-)-preS2 were confirmed by DNA sequencing to be accurately constructed as design.The luciferase activity of the pGL3-APT1 was 1.2times(P<0.01)that of the positive control plasmid pGL3-Control.And the luciferase activity of the HepG2 cells cotransfected with pcDNA3.1(-)-preS2 and pGL3-APT1 was 2.6times(P<0.01)that of the HepG2 cells cotransfected with the plasmid without preS2 gene pcDNA3.1(-)and pGL3-APT1.Conclusion The human APT1 promoter cloned in the study has high promoter activity;HBV preS2 activates human APT1 promoter.更多还原