Detection of extrahepatic hepatitis C virus replication by a novel, highly sensitive, single-tube nested polymerase chain reaction
Detection of extrahepatic hepatitis C virus replication by a novel, highly sensitive, single-tube nested polymerase chain reaction
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DOI：
10.1309/33tajlb748klmxvg
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发表时间：
2003-01-01
影响因子：
3.5
通讯作者：
Hirshfield, I
中科院分区：
文献类型：
--
作者：
Hu, Y;Shahidi, A;Hirshfield, I
我们建立了一个细胞培养系统，用于复制丙型肝炎病毒（HCV）通过使用人T和B白血病细胞系。通过使用HCV阳性合并患者血清样品体外感染这2种细胞系。在感染后的不同时间从感染的细胞系中提取HCV RNA，并分析病毒5'非翻译区的序列。丙型肝炎负链RNA检测感染的细胞系中的高度链特异性RTTH（重组嗜热栖热菌DNA聚合酶）为基础的逆转录，然后一种新的，高灵敏度，单管巢式聚合酶链反应（PCR）方法。PCR产物直接测序分析。这些结果表明HCV可以在T和B淋巴细胞中复制。这个模型应该是一个有价值的工具，详细研究的HCV复制周期的初始步骤和抗病毒分子的评价。
We established a cell culture system for the replication of hepatitis C virus (HCV) by using human T and B leukemia cell lines. These 2 cell lines were infected in vitro by using HCV-positive pooled patient serum samples. HCV RNA was extracted from infected cell lines at different times after infection, and a sequence of the virus 5' untranslated region was analyzed. Hepatitis C minus-strand RNA was detected in the infected cell lines by highly strand-specific rTth (recombinant Thermus thermophilus DNA polymerase)based reverse transcription followed by a novel, highly sensitive, single-tube nested polymerase chain reaction (PCR) method. PCR products were analyzed by direct DNA sequencing. These results indicate that the HCV can replicate in T and B lymphocytes. This model should represent a valuable tool for the detailed study of the initial steps of the HCV replication cycle and for the evaluation of antiviral molecules.