摘要
目的 乙型肝炎病毒核糖核酸酶 H (HBV- RNase H)为该病毒生活周期中负链 DNA合成所必需 ,获取该基因片段及其表达蛋白 ,为进一步的基础研究及应用研究创造条件 .方法 以 HBV全基因片段为模板 ,PCR方法定位扩增 HBV-RNase H特异性片段 ,连接于 p T7Blue克隆载体 ,酶切鉴定并测序后克隆于 p GSTag载体进行原核表达 .PAGE电泳及Western杂交鉴定表达产物 .结果 DNA测序结果显示 PCR扩增产物及载体中插入片段与已知序列相同 ;Western杂交结果表明 ,所表达产物为 GST与 RNase H的融合蛋白 .结论 HBV- RNase H基因的成功克隆及表达为进一步研究
AIM To amplify and express HBV RNase H gene fragment for further use in the studies of HBV infection diseases. METHODS The encoding gene of HBV RNase H was amplified by PCR from full length HBV gene. The amplified gene was cloned into pT7Blue vector and sequenced. The correct gene fragment was inserted into GSTag expressing vector and expressed in BL21 bacteria. The products were checked by PAGE gel and Western blot. RESULTS The nucleotide sequence was consistent with the known sequence and western blot result demonstrated that the ex pressed product was the fusion protein of GST and RNaseH. CONCLUSION The successful cloning and expression of HBV RNase H will contribute to the further research in HBV associated liver diseases.
出处
《第四军医大学学报》
2000年第3期385-387,共3页
Journal of the Fourth Military Medical University
