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基于RT-RAA和CRISPR/Cas12a的一管式玉米矮花叶病毒快速检测方法

A one-tube rapid detection method for maize dwarf mosaic virus based on RT-RAA and CRISPR/Cas12a

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【作者】 侯雨萌赵振兴王思元董铮胡中泽周涛张永江

【Author】 HOU Yumeng;ZHAO Zhenxing;WANG Siyuan;DONG Zheng;HU Zhongze;ZHOU Tao;ZHANG Yongjiang;Chinese Academy of Inspection and Quarantine;College of Plant Protection, China Agricultural University;Taizhou Institute of Agricultural Sciences, Jiangsu Academy of Agricultural Sciences;

【通讯作者】 周涛;张永江;

【机构】 中国检验检疫科学研究院中国农业大学植物保护学院江苏省农业科学院泰州农业科学研究所

【摘要】 玉米矮花叶病毒(Maize dwarf mosaic virus, MDMV)是一种重要的检疫性病毒,严重影响玉米的生产。为了有效防治玉米矮花叶病,本研究基于反转录重组酶介导等温核酸扩增(RT-RAA)技术和CRISPR/Cas12a系统构建了一种MDMV快速检测方法,将所有试剂集中在一个试管中进行反应,并通过侧向流动试纸条(LFD)检测反应结果。本研究筛选得到的最佳反应条件为,引物添加量2.8μL、FB报告分子浓度100 nmol/L、RT-RAA系统反应时间20 min、CRISPR/Cas12a系统反应时间20 min。本研究构建的基于RT-RAA和CRISPR/Cas12a的一管式玉米矮花叶病毒快速检测方法特异性强、灵敏度高,且所用试验材料方便携带和运输,适用于现场检测。

【Abstract】 Maize dwarf mosaic virus(MDMV) is an important quarantine virus that seriously affects maize production. To effectively prevent and control maize dwarf mosaic disease, this study constructed a rapid detection method for MDMV based on reverse transcription-recombinase aided amplification(RT-RAA) technology and the CRISPR/Cas12a system. All reagents were reacted in a single test tube, and the reaction results were detected by lateral flow dipstick(LFD). The optimal reaction conditions screened in this study were as follows: primer addition amount of 2.8 μL, FB reporter molecule concentration of 100 mol/L, RT-RAA system reaction time of 20 minutes, and CRISPR/Cas12a system reaction time of 20 minutes. The one-tube rapid detection method for maize dwarf mosaic virus based on RT-RAA and CRISPR/Cas12a constructed in this study has strong specificity, high sensitivity, and the experimental materials used are convenient for carrying and transportation, making it suitable for on-site detection.

【基金】 国家重点研发计划项目(2021YFD1400100、2021YFD1400103)
  • 【文献出处】 江苏农业学报 ,Jiangsu Journal of Agricultural Sciences , 编辑部邮箱 ,2025年02期
  • 【分类号】S435.131
  • 【下载频次】46
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