An improved cell-free system for picornavirus synthesis
An improved cell-free system for picornavirus synthesis
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DOI：
10.1016/j.jviromet.2007.01.026
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发表时间：
2007-06-01
影响因子：
3.1
通讯作者：
Imataka, Hiroaki
中科院分区：
文献类型：
--
作者：
Kobayashi, Tominari;Mikami, Satoshi;Imataka, Hiroaki
感染性病毒的无细胞合成是阐明病毒复制机制和筛选抗病毒药物的理想工具。在本研究中，通过采用透析系统与核酶技术相结合，增强了从 HeLa 和 293-F 细胞提取物中的 RNA 合成脑心肌炎病毒 (EMCV) 的能力。尽管透析系统并未大大加速 EMCV 多蛋白的翻译和加工，但透析显着增强了病毒 RNA 的从头合成，导致合成的 EMCV 滴度比传统批量系统增加了八倍以上。此外，在 5' 端具有锤头状核酶序列的合成 EMCV RNA 可作为透析系统中病毒合成的有效模板。因此，该系统为EMCV体外突变分析提供了机会。 (c) 2007 Elsevier B.V. 保留所有权利。
Cell-free synthesis of an infectious virus is an ideal tool for elucidating the mechanism of viral replication and for screening anti-viral drugs. In the present study, the synthesis of Encephalomyocarditis virus (EMCV) from its RNA in HeLa and 293-F cell extracts was enhanced by employing a dialysis system in combination with a ribozyme technology. Although translation and processing of the EMCV polyprotein were not accelerated greatly by the dialysis system, de novo synthesis of viral RNA was enhanced considerably by dialysis, leading to a greater than eight-fold increased titer of synthesized EMCV compared with a conventional batch system. Furthermore, a synthetic EMCV RNA with a hammerhead ribozyme sequence at its 5'-end served as an efficient template for viral synthesis in the dialysis system. Therefore, this system provides opportunities for mutational analyses of EMCV in vitro. (c) 2007 Elsevier B.V. All rights reserved.