构建双子病毒传染性克隆,用于在植物中进行农业化
Ivair J Morais1, Alice K Inoue-Nagata2, Erich Y T Nakasu3
1Department of Phytopathology, University of Brasília, Brasília, DF, Brazil.
Methods in molecular biology (Clifton, N.J.)
|November 21, 2023
概括
本研究详细介绍了创建传染性双胞胎病毒克隆的方法,这对于标准化宿主病毒研究至关重要. 介绍了两种协议,限制酶消化和吉布森组合,用于构建二元克隆,以确保全长后代DNA生产.
科学领域:
- 植物病毒学 植物病毒学
- 分子生物学分子生物学
- 分子克隆是一种分子克隆.
背景情况:
- 双胞胎病毒具有单链DNA基因组 (单片或双片),对于宿主病毒相互作用研究至关重要.
- 标准化的双子病毒注射剂对于可重现的实验结果至关重要.
- 生产全长后代DNA需要特定的基因组配置在传染性克隆.
研究的目的:
- 描述一种用于制备传染性双胞胎病毒克隆的综合性协议.
- 介绍两种不同的方法来构建二维双胞胎病毒克隆:限制酶消化/结合和吉布森组合.
- 为了促进用于双子病毒研究的标准化注射器生产.
主要方法:
- 从受双胞胎病毒感染的宿主中提取DNA.
- 选择适当的限制性内核酶用于克隆.
- 使用限制性内核酶消化和结合,构建二元传染性克隆.
- 使用吉布森组件构建二元传染性克隆.
主要成果:
- 提供了准备传染性双胞胎病毒克隆的详细方案.
- 限制酶消化/结合和吉布森组合都对构建二次克隆有效.
- 描述的方法确保了后代DNA生产复制起源的多个副本的存在.
结论:
- 描述的协议使得感染性双胞胎病毒克隆的标准化生产成为可能.
- 这些方法对于在双子病毒研究中推进宿主病毒相互作用研究至关重要.
- 可靠的传染性克隆的开发支持可再生植物病毒研究.
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