论文部分内容阅读
构建具有不同类型和长度的5’非编码区(UTR)及3’UTR的丙型肝炎病毒(HCV)全长cDNA克隆,体外转录制备RNA,转染肝癌细胞HepG2,研究不同RNA转录体在细胞内的复制情况。通过RT-PCR与Westernblot等方法证明:蛋白编码区来源于1a基因型并具有1b型5’UTR及3’UTR(全长或缺失98nt保守区)的2种嵌合型HCVRNA,都可以在病毒易感细胞中复制和表达;而以脑心肌炎病毒(EMCV)内核糖体进入位点(IRES)替代HCV5’UTR的嵌合型RNA转染细胞,检测不到病毒正、负链RNA与蛋白表达。以上结果说明HCV1b型5’UTR与3’UTR可以支持1a型病毒RNA的正常复制,3’UTR保守区的存在与否,对病毒复制影响不大。
To construct full-length Hepatitis C virus (HCV) cDNA clones with different types and lengths of 5 ’untranslated region (UTR) and 3’UTR, RNA was prepared and transfected into HepG2 hepatocellular carcinoma cells in vitro to study the effect of different RNA transcripts Within the copy. The results of RT-PCR and Westernblot showed that two kinds of chimeric HCVRNA, which are derived from genotype 1a and have type 1b 5’UTR and 3’UTR (full-length or deletion of 98nt conserved region) While the chimeric RNA transfected cells with HCV 5 ’UTR were replaced by the internal ribosomal entry site (IRES) of encephalomyocarditis virus (EMCV), and no positive and negative RNA and protein expression of the virus were detected . The above results indicate that HCV1b 5’UTR and 3’UTR can support the normal replication of type 1a viral RNA, and the presence or absence of 3’UTR conserved region has little effect on virus replication.