进境苹果中3种检疫性真菌三重PCR同步检测方法的建立  

Establishment of triplex PCR synchronous detection methods for three quarantine fungi in imported apples

作　　者：田沂民 张露茜 葛立傲 焦彬彬 胡培龙 徐飞 于翠 杨翠云 Tian Yimin;Zhang Luxi;Ge Liao;Jiao Binbin;Hu Peilong;Xu Fei;Yu Cui;Yang Cuiyun(Shanghai Customs,Shanghai 200135,China;School Affiliated to Jinyuan Senior High School;Shanghai Jinshan District Vegetable Technology Promotion Center)

机构地区：[1]上海海关,上海200135 [2]上海市晋元高级中学附属学校 [3]上海市金山区蔬菜技术推广中心

出　　处：《植物检疫》2018年第5期18-23,共6页Plant Quarantine

基　　金：上海出入境检验检疫局科技计划项目(HK001-2017)

摘　　要：苹果壳色单隔孢溃疡病菌(Botryosphaeria stevensii)、美澳型核果褐腐病菌(Monilinia fructicola)和丁香疫霉病菌(Phytophthora syringae)是为害多种水果和其他植物的重要检疫性真菌,均能侵染苹果果实导致烂果,并造成严重的经济损失。本研究设计和筛选了3对特异性引物,建立了同步检测3种检疫性真菌的普通PCR和三重PCR方法。设计的引物Bs-368 F/R、M.cola 986F/R和Psy1/Psy2能分别特异性地扩增出B.stevensii、M.fructicola和P.syringae所有供试菌株的DNA,而对相应多个近似种的DNA则无扩增。灵敏度试验结果表明,上述3对引物分别检测B.stevensii、M.fructicola和P.syringae DNA的灵敏度是41.7 pg、47.4 pg和0.375 pg,三重PCR同时检测到3种病菌DNA的灵敏度是4.44 ng。该方法可以满足对进境苹果果实携带的苹果壳色单隔孢溃疡病菌、美澳型核果褐腐病菌和丁香疫霉病菌的快速初筛,加速进境新鲜水果的快检快放。Botryosphaeria stevensii,Monilinia fructicola and Phytophthora syringae are important quarantine fungi of China. They are harmful to a variety of fruits and other plants leading to serious economic loss. In this study,three pairs of specific primers were designed and screened,furthemore common PCR and triple PCR methods for simultaneous detection of three kinds of quarantine fungi were established.The designed primers Bs-368 F/R、M.cola 986 F/R and Psy1/Psy2 could respectively specifically amplify all strains DNA of B. stevensii,M. fructicola and P. syringae,but could not amplify the strains DNA of closely related species. Sensitivity test results showed that the sensitivity of the above three pairs of primers was respectively 41.7 pg,47.4 pg,0.375 pg, and the triple PCR was able to detect the sensitivity of the three DNA mixtures to 4.44 ng. This method can meet the rapid and effective screening purpose of Botryosphaeria stevensii,Monilinia fructicola and Phytophthora syringae in imported apples,and speed up the entry of fresh fruit.

关 键 词：苹果壳色单隔孢溃疡病菌 美澳型核果褐腐病菌 丁香疫霉病菌 三重PCR 同步检测 

分 类 号：S41[农业科学—植物保护]