实时荧光PCR检测瓜炭疽病菌  

Detection of Colletotrichum orbiculare by the real-time PCR

作　　者：王翀[1] 张小菊[1] 张祥林[1] 张伟[1] 张瑜[1] 李亚伟[1] 孙燕飞[2] 

机构地区：[1]新疆出入境检验检疫局,乌鲁木齐830063 [2]石河子大学,石河子832000

出　　处：《植物保护》2016年第2期129-135,共7页Plant Protection

基　　金：国家质量监督检验检疫总局科研计划项目(2013IK287;2011IK168);国家科技部质检公益性行业科研专项(201310091)

摘　　要：采用实时荧光PCR技术建立了瓜炭疽病菌(Colletotrichum orbiculare)的检测方法。根据瓜炭疽病菌甘油醛-3-磷酸脱氢酶(GAPDH)基因和谷氨酰胺合成酶(GS)基因序列,设计了该病菌特异性引物和TaqMan探针,并对所设计的引物和探针的反应条件进行了优化。采用本试验建立的实时荧光PCR方法对瓜上的其他菌株及近似菌株进行检测,可将瓜炭疽病菌与其他病原菌区分开。灵敏度试验表明,25μL体系中只要有39.6pg的核酸量就可以被检测到,检测灵敏度达到1.584pg/μL,比普通PCR检测灵敏度高100倍。同时对田间采集的病株和未知样品进行的检测证明了引物和TaqMan探针的特异性。The detection of Colletotrichum orbiculare by real-time PCR was established.The specific primers and TaqMan probes were designed according to glyceraldehyde-3-phosphate dehydrogenase（GAPDH）gene and glutamine synthase（GS）gene sequences in C.orbiculare,and the reaction conditions were optimized.C.orbiculare could be detected from other strains in melon and other similar strains by real-time PCR.The sensitivity test indicated that 39.6 pg DNA could be detected in25μL reaction system.The real-time PCR sensitivity could reach to1.584 pg/μL,100 times more sensitive than ordinary PCR.The primers and Taqman probe were shown to be specific by detecting infected plants sampled in the field and unknown samples.

关 键 词：实时荧光PCR 检测 瓜炭疽病菌 

分 类 号：S436.5[农业科学—农业昆虫与害虫防治]