实时荧光定量PCR检测核桃胶胞炭疽菌  

Detecting Colletotrichum gloeosporioides in Walnut by Real-time PCR

作　　者：余鹏举 朱天辉[1] 万雪琴[1] 李姝江 王若梅 韩珊[1] Yu Pengju;Zhu Tianhui;Wan Xueqin;Li Shujiang;Wang Ruomei;Han Shan(Sichuan Agricultural University,Chengdu 611130,P.R.China)

机构地区：[1]四川农业大学,成都611130

出　　处：《东北林业大学学报》2018年第11期87-90,100,共5页Journal of Northeast Forestry University

摘　　要：为快速灵敏检测核桃炭疽病病原菌,根据Gen Bank中炭疽属(Colletotrichum)不同种的ITS序列差异,设计了核桃炭疽病病原菌胶胞炭疽菌(C. gloeosporioides)的特异性引物YH1/YH2,并优化建立了检测核桃胶胞炭疽菌的SYBR Green I实时荧光定量PCR检测体系。结果表明:该引物能从实验室分离保存鉴定的13株核桃胶胞炭疽菌中特异性扩增出306bp的目的条带,而对于其他6种核桃常见病害病原菌、健康核桃植株组织及所有微生物总基因组DNA均未扩增出特异性条带;建立的实时荧光定量PCR检测体系灵敏度为1×10-3mg·L-1,是常规PCR检测方法的100倍;对10份采集于四川、陕西、湖北等地的林间感病核桃植株组织进行定量检测,实时荧光定量PCR的阳性检出率为80%,且样品病症表现越明显检测到的含菌量越高。建立的SYBR Green I实时荧光定量PCR检测方法可用于核桃感病组织内胶胞炭疽菌的分子鉴定和早期检测。For the rapid and sensitive detection of pathogenic fungi of walnut anthracnose,a pair of specific primers YH1/YH2 was designed to detect walnut Colletotrichum gloeosporioides according to the ITS sequence of different species in GenBank.The results showed that 13 strains of C.gloeosporioides were amplified with 306 bp bands,while no specific bands were amplified for the pathogens from other six walnut common diseases.The sensitivity of the established SYBR Green I real-time PCR detection system was 1×10-3 mg·L-1,which was 100 times higher than that of the conventional PCR detection method.The samples were collected in Sichuan,Shaanxi and Hubei Province,etc.The positive rate of the real-time PCR was 80%,and the more obvious the disease was detected,the higher the concentration of pathogenic fungi was detected.The established SYBR Green I real-time PCR method can be used for molecular identification and early detection of C.gloeosporioides in tissues of walnut.

关 键 词：核桃 胶胞炭疽菌 SYBR Green I实时荧光定量PCR 分子检测 

分 类 号：S763.1[农业科学—森林保护学] S792.13[农业科学—林学]