HBVX蛋白真核表达质粒的构建及其对反式转录激活作用的影响  

Construction of HBV X mammalian expression plasmid and detection of transcriptional activity of HBV X protein

作　　者：杨益大[1] 郑临[1] 杨芊[1] 马亦林[1] 

机构地区：[1]浙江大学医学院附属第一医院传染病科,杭州310003

出　　处：《中华传染病杂志》2002年第4期222-224,共3页Chinese Journal of Infectious Diseases

基　　金：浙江省自然科学基金资助项目 ( 4 910 10 )

摘　　要：目的　构建HBVX蛋白真核表达质粒并了解其对反式转录激活作用的影响。方法　用亚克隆方法构建HBVX蛋白真核表达质粒 ,以氯霉素乙酰转移酶转化率分析 (CAT试验 )揭示HBX蛋白的反式转录激活作用。结果　HBVX蛋白真核表达质粒转染HepaG2细胞后 ,以WesternBlot检测发现 2 μg、5 μg和 10 μg的 p5SGUTPLHBXDNA均能有效表达 ;CAT试验揭示 2 μg、5 μg、10μg的 p5SGUTPLHBXDNA对报道基因 pHE2×CAT的14 C -标记乙酰化氯霉素转化率分别为 (4 5 .5± 2 5 .9) %、(6 5 .6± 14.4) %和 (6 8.8± 19.7) %。Objective To construct HBV X mammalian expression plasmid and establishment the method to detect transcriptional activity of HBV X protein. Methods HBV X mammalian expression plasmid pSG5UTPLHBX was constructed using subclone method. Choramphenicol Acetyltransferase Assay(CAT) was set up to detect the transcriptional transactivation of HBX protein in vitro. Results Transfected in HepaG2 cells, all 2 μg, 5 μg, 10 μg of pSG5UTPLHBX DNA were well expressed. When 2 μg, 5 μg, 10 μg of pSG5UTPLHBX DNA were transfected together with reporter gene pHEC 2×CAT in the HepaG2 cells, the CAT activities were (45.5 ±25.9)%, (65.6±14.4)%, and (68.8 ±19.7)%, respectively.Conclusions These results lay a fundation for further study of transcriptional transactivation of HBX protein.

关 键 词：HBV X蛋白 真核表达 质粒 乙型肝炎 反式转录激活 

分 类 号：R346[医药卫生—基础医学]