INVESTIGATION OF REPLICATION MECHANISM AND PATHOGENESIS OF PARAMYXOVIRUS BY USING INFECTIOUS cDNA CLONE

利用感染性cDNA克隆研究副粘病毒的复制机制和发病机制

• 批准号: 04454200
• 负责人: ITO Yasuhiko
• 金额: $ 4.29万
• 依托单位: MIE UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454200/
• 关键词: paramyxovirus; parainfluenza virus; infectious cDNA; RNA Replicon; HPIV-2; 発現ベクター

At first, the nucleotide sequences of the L gene of Paramyxovirus, simian viruses 5 and 41 were determined. Therefore, amino acid sequences of components of replicative complex, NP, P, and L proteins were also determined. Full-length cDNA clones for NP,P, and L protein could be obtained. Next, artificial mini-genomes, Leader sequence-R1-reporter gene-R2-Trailer sequence, were constructed. The mini-genomes were transfected into human parainfluenza type 2 virus or simian virus 41-infected cells. In this system, replication, transcription and RNA editing were occurred. However, the efficiency was not so high. Subsequently, the mini-genomes, cDNA for NP, P, and L proteins were transfected into the cells infected recombinant vaccinia virus containing T7 polymerase. In this new system, high efficiency of transcription, replication and RNA editing were observed. In another experiment, full-length cDNA clone for human parainfluenza virus type 2 was nearly constructed.

首先，测定了副粘病毒、猿猴病毒5号和41号的L基因的核苷酸序列。因此，还确定了复制复合体、NP、P和L蛋白的组分的氨基酸序列。可以获得NP、P和L蛋白的全长cDNA克隆。接下来，构建了人工迷你基因组，Leader序列-R1-报告基因-R2-Trailer序列。将微型基因组转染到人类副流感 2 型病毒或猿猴病毒 41 感染的细胞中。在这个系统中，发生了复制、转录和RNA编辑。不过，效率并没有那么高。随后，将小基因组、NP、P和L蛋白的cDNA转染到感染含有T7聚合酶的重组痘苗病毒的细胞中。在这个新系统中，观察到转录、复制和 RNA 编辑的高效率。在另一项实验中，人类副流感病毒2型的全长cDNA克隆已基本构建完成。

项目成果

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