马铃薯病毒 Y 的核包涵蛋白 B 驱动体外复制系统

IF 2.3 3区 农林科学 Q1 AGRONOMY
Jiayu Fu, Zichen Li, Minjun Liu, Guowei Geng
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引用次数: 0

摘要

马铃薯病毒 Y(PVY)是影响全球马铃薯和其他作物的重要病原体。目前对 PVY 的研究主要集中在菌株分析和蛋白质与宿主的相互作用方面。然而,对 PVY 体外复制的研究仍然有限,主要原因是缺乏专门的 PVY 体外复制系统。本研究旨在开发一种由 PVY 的核包涵蛋白 B(NIb)介导的体外复制系统。通过 IPTG 梯度实验,我们确定 MBP-NIb 的最佳表达浓度为 0.2 mM。结果表明,在大肠杆菌中表达 NIb 的最佳温度为 37 °C,溶解度约为 10%,这足以满足随后使用 MBP 标签进行亲和层析的需要。纯化的 MBP-NIb 能在体外特异性识别 PVY 阳性有义 RNA 的 3′ 端区和阴性有义 RNA 的 5′ 端区,促进病毒 RNA 互补链的合成。据我们所知,这是首次成功建立 PVY 体外复制系统。该系统将推动对 PVY 复制机制的研究。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Nuclear Inclusion Protein B-Driven In Vitro Replication System for Potato Virus Y

Nuclear Inclusion Protein B-Driven In Vitro Replication System for Potato Virus Y

Potato virus Y (PVY) is a significant pathogen affecting potato and other crops globally. Current research on PVY primarily concentrates on strain analysis and protein interactions with the host. However, studies on PVY replication in vitro remain limited, primarily due to the absence of a dedicated in vitro replication system for PVY. This study aimed to develop an in vitro replication system mediated by the nuclear inclusion protein B (NIb) of PVY. Through IPTG gradient experiments, we determined that the optimal expression concentration of MBP-NIb is 0.2 mM. The results indicate that the optimal temperature for NIb expression in Escherichia coli is 37 °C, yielding approximately 10% solubility, which is sufficient for subsequent affinity chromatography using the MBP tag. The purified MBP-NIb specifically recognizes the 3′ terminal region of the positive-sense RNA and the 5′ terminal region of the negative-sense RNA of PVY in vitro, facilitating the synthesis of complementary strands of the viral RNAs. To our knowledge, this is the first successful establishment of an in vitro replication system for PVY. This system will advance the study of PVY replication mechanisms.

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来源期刊
Potato Research
Potato Research AGRONOMY-
CiteScore
5.50
自引率
6.90%
发文量
66
审稿时长
>12 weeks
期刊介绍: Potato Research, the journal of the European Association for Potato Research (EAPR), promotes the exchange of information on all aspects of this fast-evolving global industry. It offers the latest developments in innovative research to scientists active in potato research. The journal includes authoritative coverage of new scientific developments, publishing original research and review papers on such topics as: Molecular sciences; Breeding; Physiology; Pathology; Nematology; Virology; Agronomy; Engineering and Utilization.
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