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新型鸭呼肠孤病毒感染性分子克隆的构建与鉴定。

Construction and characterization of an infectious molecular clone of novel duck reovirus.

机构信息

National Engineering Research Center for Poultry, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, No. 518 Ziyue Road, Minhang District, Shanghai 200241, PR China.

出版信息

J Gen Virol. 2018 Apr;99(4):449-456. doi: 10.1099/jgv.0.001036. Epub 2018 Feb 26.

Abstract

Novel duck reovirus (NDRV), the prototype strain of the species Avian orthoreovirus (ARV), is currently an infectious agent for ducks. Studies on NDRV replication and pathogenesis have been hampered by the lack of an available reverse-genetics system. In this study, a plasmid-based reverse-genetics system that is free of helper viruses has been developed. In this system, 10 full-length gene segments of wild-type NDRV TH11 strain are transfected into BSR-T7/5 cells that express bacteriophage T7 RNA polymerase. Production of infectious virus was shown by the inoculation of cell lysate derived from transfected cells into 10-day-old duck embryos. The in vivo growth kinetics and infectivity of the recombinant strains were identical to those of the wild-type strain. These viruses grew well and were genetically stable both in vitro and in vivo. Altogether, these results show the successful production of an infectious clone for NDRV. The infectious clone reported will be further used to elucidate the mechanisms of host tropism, viral replication and pathogenesis, as well as immunological changes induced by NDRV.

摘要

新型鸭呼肠孤病毒(NDRV)是禽呼肠孤病毒(ARV)种的原型株,目前是鸭的一种感染性病原体。由于缺乏可用的反向遗传学系统,对 NDRV 复制和发病机制的研究受到了阻碍。本研究开发了一种无辅助病毒的基于质粒的反向遗传学系统。在该系统中,10 个全长的野生型 NDRV TH11 株基因片段被转染到表达噬菌体 T7 RNA 聚合酶的 BSR-T7/5 细胞中。通过将转染细胞的细胞裂解物接种到 10 日龄鸭胚中,证明了感染性病毒的产生。重组株的体内生长动力学和感染力与野生型株相同。这些病毒在体外和体内均能良好生长且遗传稳定。总之,这些结果表明成功生产了 NDRV 的感染性克隆。所报道的感染性克隆将进一步用于阐明宿主嗜性、病毒复制和发病机制以及 NDRV 诱导的免疫变化的机制。

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