临床肝胆病杂志2011，Vol.27Issue(1)：57-60,4.
携带IFN-β基因的丙型肝炎病毒微型基因组构建及抑制病毒复制的效应
Construction of HCV minigenome containning IFN-β and investigation of its inhibitory effect on viral replication
摘要
Abstract
Objective To construct minigenome containning IFN-β responsive to HCV and to study its inhibitory effect on viral replication. Methods Total RNA was extracted from lymphocytes stimulated by poly I:C and was reversely transcripted into cDNA.IFN-β gene was amplified by PCR and was cloned into pT7-5U△131-315rI3Urz in antisense direction to obtain pT7-5U△131-315rIFNI3Urz.The minigenome 5U△131-315rIFNI3URz was amplified and inserted into the plasmid pcDNA3.1 by positioning the minigenome immediately adjacent to the transcription start site of the cytomegalovirus (CMV) promoter, resulting in pCMV-5U△131-315rIFNI3URz.The pT7-SU△131-315rIFNI3Urz was transcripted into RNA in vitro, then transfected into Huh7.SBB7. After 48h, cell was harvested and total proteins were extracted for Western Blot, IFN-β antibody as the first antibody. When the pCMV-SUrIFNI3URz was transfected into Huh7.SJFH-1 cell the real-time PCR assay was used to detect HCV RNA replication level. Results The Huh7.5-JFH-I cell line which was developed by transfecting infectious RNA into Huh7.5 cell which can replicate autonomously and virus particle can be packed successfully. IFN-β could be specifically expressed after 5UrIFNI3Urz RNA and pCMV-SUrIFNI3URz was trasnfected into Huh7.SBB7 cell lines. The level of HCV RNA replication was distinctly decreased when Huh7.5-JFH-1 cell line was transfected with pCMV-SUrIFNI3URz. Conclusion The minigenome containning inverted IFN-β could specifically be expressed in HCV infected.cell lines, the viral replication level was decreased to a certain extent. This study provides the new idea of anti-HCV infection.关键词
肝炎病毒属/基因组,病毒/干扰素β/复制子Key words
hepacivirus/ genome, viral/ interferonc-beta/ replicon分类
医药卫生引用本文复制引用
杨敬,雷迎峰,尹文,吕欣,陈任安,安群星,姚敏,贾战生,徐志凯..携带IFN-β基因的丙型肝炎病毒微型基因组构建及抑制病毒复制的效应[J].临床肝胆病杂志,2011,27(1):57-60,4.基金项目
国家自然科学基金资助项目(30700710,30700030,30600564) （30700710,30700030,30600564）
国家"863"专项课题资助项目(2007AA02Z441) （2007AA02Z441）
