https://www.ahs.ac.cn/images/0513-353X/images/top-banner1.jpg|#|苹果
https://www.ahs.ac.cn/images/0513-353X/images/top-banner2.jpg|#|甘蓝
https://www.ahs.ac.cn/images/0513-353X/images/top-banner3.jpg|#|菊花
https://www.ahs.ac.cn/images/0513-353X/images/top-banner4.jpg|#|灵芝
https://www.ahs.ac.cn/images/0513-353X/images/top-banner5.jpg|#|桃
https://www.ahs.ac.cn/images/0513-353X/images/top-banner6.jpg|#|黄瓜
https://www.ahs.ac.cn/images/0513-353X/images/top-banner7.jpg|#|蝴蝶兰
https://www.ahs.ac.cn/images/0513-353X/images/top-banner8.jpg|#|樱桃
https://www.ahs.ac.cn/images/0513-353X/images/top-banner9.jpg|#|观赏荷花
https://www.ahs.ac.cn/images/0513-353X/images/top-banner10.jpg|#|菊花
https://www.ahs.ac.cn/images/0513-353X/images/top-banner11.jpg|#|月季
https://www.ahs.ac.cn/images/0513-353X/images/top-banner12.jpg|#|菊花

园艺学报 ›› 2019, Vol. 46 ›› Issue (7): 1409-1416.doi: 10.16420/j.issn.0513-353x.2018-0840

• 新技术与新方法 • 上一篇    下一篇

柑橘鳞皮病毒RT-LAMP检测方法的建立与应用

李 敏,周天宇,吴佳星,周 彦,曹孟籍*,李中安*   

  1. 西南大学,中国农业科学院柑桔研究所,重庆 400712
  • 出版日期:2019-07-25 发布日期:2019-07-25
  • 基金资助:
    国家重点研发计划政府间国际科技创新合作重点专项(2017YFE0110900);重庆市基础科学与前沿技术研究专项(cstc2017jcyjBX0016);重庆市研究生科研创新项目(CYS18133,CYS17086)

Establishment and Application of Loop-mediated Isothermal Amplification on Citrus psorosis virus

LI Min,ZHOU Tianyu,WU Jiaxing,ZHOU Yan,CAO Mengji*,and LI Zhongan*   

  1. Citrus Research Institute,Southwest University,Chinese Academy of Agricultural Sciences,Chongqing 400712,China
  • Online:2019-07-25 Published:2019-07-25

摘要: 根据NCBI数据库柑橘鳞皮病毒(Citrus psorosis virus,CPsV)的外壳蛋白(coat protein,CP)基因序列设计了4组特异性引物,通过一系列优化,筛选并获得1组特异性引物。以感染CPsV的柑橘叶片总RNA为模板,采用一步法RT-LAMP(One-step reverse transcription loop-mediated isothermal amplification),在60 ℃下反应60 min,对扩增产物进行琼脂糖电泳分析,同时测定其特异性和灵敏度。结果表明,建立的RT-LAMP检测方法能特异性扩增CPsV,灵敏度为RT-PCR的10倍。用RT-LAMP 方法对87个田间样品进行检测,发现有5个样品感染了CPsV,其检测结果与RT-PCR法一致。RT-LAMP法具有特异性强、灵敏度高、操作简单、快速等特点,适合对CPsV样品的快速检测与鉴定。

关键词: 柑橘, 柑橘鳞皮病毒, RT-LAMP, 检测

Abstract: Four sets of specific primer pairs in the conserved region of the CP(coat protein)gene of Citrus psorosis virus(CPsV)were designed for One-step RT-LAMP detection system based on NCBI database. One feasible pair of primers was screened to be suitable for this system. Template RNA extracted from CPsV infected citrus leaves was used for One-step RT-LAMP,and isothermal conditions were optimized to be 60 ℃ for 60 minutes. The agarose gel electrophoresis analysis of RT-LAMP products indicated that the established RT-LAMP system was specific and sensitive in detecting CPsV and 10-fold higher than RT-PCR method in sensitivity. Of 87 samples detected by this RT-LAMP system,five were found to be infected by CPsV,showing the detection result was consistent with RT-PCR method. Hence this RT-LAMP system is applicable for rapid and accurate detection and identification of CPsV.

Key words: citrus, Citrus psorosis virus(CPsV), RT-LAMP, detection

中图分类号: