基于TaqMan MGB探针的菊花花枯病菌实时荧光PCR检测方法  

TaqMan MGB-based real-time PCR method for the detection of Stagonosporopsis chrysanthemi

作　　者：滕少娜 孙涛 孔德英 马冠华[2] 何玲 谢鑫[3] Theng Shaona;Sun Tao;Kong Deying;Ma guanhua;He Ling;Xie Xin(Technology Center of Chongqing Customs District,Chongqing 400020,China;College of Plant Protection,Southwest University;College of Agriculture,Guizhou University)

机构地区：[1]重庆海关技术中心,重庆400020 [2]西南大学植物保护学院 [3]贵州大学农学院

出　　处：《植物检疫》2024年第4期39-44,共6页Plant Quarantine

基　　金：原国家质检总局项目(2017IK207);重庆市科技局项目(cstc2017shms-xdny0145&cstc2019jscx-msxmX0410)。

摘　　要：菊花花枯病菌已被列入我国进境植物检疫性有害生物名录。目前,随着真菌分类系统的调整以及“一菌一名”原则的实施,菊花花枯病菌及其近似种分类地位已经发生了重大调整。本研究根据菊花花枯病菌模式菌株与其近似种ITS序列的差异,设计了可特异性检测菊花花枯病菌的引物SL1/SL2和探针Probe-M,建立了菊花花枯病菌的实时荧光PCR检测方法。特异性试验结果表明,供试菌株中仅菊花花枯病菌有阳性扩增,而其他供试菌株及空白对照均无荧光信号增加。灵敏度试验结果表明,在20μL反应体系内,检测灵敏度可达到26 fg/μL DNA。该方法特异性强、灵敏度高、检测速度快,整个检测过程在1.5 h内完成,满足了口岸对菊花花枯病菌快速检疫鉴定的需求。The pathogen Stagonosporopsis chrysanthemi has been included in the list of imported plant quarantine pests in China. At present,with the adjustment of the classification system of fungi and the implementation of the principle of "one fungus,one name",the classification status of S. chrysanthemi and related species have been greatly adjusted. In this study,a pair of specific primers SL1/SL2 and Probe-M according to the difference of ITS sequence between model strain of S. chrysanthemi and its related species were designed and synthesized for real-time fluorescent PCR assay. The specific test results showed that only S. chrysanthemi had positive amplification,while other strains and blank controls had no increased fluorescence signal. The sensitivity test results showed that the detection limit of this method was 26 fg/μL DNA in 20 μL reaction mixture. This method exhibits strong specificity,high sensitivity,and rapid detection speed. The entire detection process is completed within 1.5 hours,meeting the requirements for rapid quarantine and identification of S. chrysanthemi at ports.

关 键 词：菊花花枯病菌 ITS序列 TAQMAN-MGB探针 实时荧光PCR 

分 类 号：S432.1[农业科学—植物病理学]