Huang Yao-Wei, Li Long, Yu Lian
Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou 310029, China.
Sheng Wu Gong Cheng Xue Bao. 2004 May;20(3):311-8.
The recovery of the virus from genetic materials in in vitro culture systems or sensitive animals is called virus rescue. A functional infectious clone of RNA virus provides unlimited possibility for genetic studies and the related reverse genetics system that allows directed genetic manipulation of an RNA virus is an extremely powerful research tool. In the past twenty years, especially since the first infectious clone of a negative-stranded RNA virus was reported in the mid-1990's, the reverse genetics systems have been available for nearly all the major human and animal RNA virus groups. The article reviews the progress of this technology, highlighting the obstacles in the construction of reverse genetics systems for major groups of human as well as animal RNA viruses and how the virologists overcame them. There are mainly four external expression systems for construction of the RNA virus reverse genetics systems basing on the kind of RNA viruses. These systems include in vitro RNA transcripts, RNA polymerase I-driven expression plasmids, RNA polymerase II-driven expression plasmids and modified vaccinia virus/T7 RNA polymerase-driven expression system. In particular, the viral nucleoprotein and polymerase proteins are required to assemble the viral ribonucleoprotein (RNP) complexes for the rescue of the negative-stranded RNA viruses. Relevant topics about the rescue of the typical viruses are discussed, including poliovirus with the de novo synthesis, Coronaviridae with the largest size of genome, Flaviviridae with the instable clones, HCV with the quasispecies nature, nodaviruses with the virus-host interaction, influenza virus with the RNA pol I transcription system, Arenavirdae with the ambisense coding strategies etc.
在体外培养系统或敏感动物中从遗传物质中拯救出病毒被称为病毒拯救。RNA病毒的功能性感染性克隆为遗传学研究提供了无限可能,而允许对RNA病毒进行定向基因操作的相关反向遗传学系统是一种极其强大的研究工具。在过去二十年中,特别是自20世纪90年代中期报道了第一个负链RNA病毒的感染性克隆以来,几乎所有主要的人类和动物RNA病毒组都已建立了反向遗传学系统。本文综述了该技术的进展,重点介绍了构建人类和动物主要RNA病毒组反向遗传学系统时遇到的障碍以及病毒学家是如何克服这些障碍的。基于RNA病毒的种类,构建RNA病毒反向遗传学系统主要有四种外源表达系统。这些系统包括体外RNA转录本、RNA聚合酶I驱动的表达质粒、RNA聚合酶II驱动的表达质粒以及改良痘苗病毒/T7 RNA聚合酶驱动的表达系统。特别是,负链RNA病毒的拯救需要病毒核蛋白和聚合酶蛋白来组装病毒核糖核蛋白(RNP)复合物。文中还讨论了典型病毒拯救的相关话题,包括从头合成的脊髓灰质炎病毒、基因组最大的冠状病毒科、克隆不稳定的黄病毒科、具有准种性质的丙型肝炎病毒、具有病毒-宿主相互作用的诺达病毒、具有RNA pol I转录系统的流感病毒、具有双义编码策略的沙粒病毒科等。