刺盘孢属真菌PCR检测方法的建立

Establishment of a PCR Test Method for Detecting Colletotrichum

【目的】刺盘孢属(Colletotrichum)真菌是重要的植物病原菌,引起植物炭疽病。建立刺盘孢属真菌PCR检测方法。【方法】比对分析刺盘孢属及其近似属的ITS序列,设计刺盘孢属特异性引物,建立PCR扩增体系,验证引物的特异性和灵敏度。【结果】设计出刺盘孢属特异性引物ITS-c3/c4,建立了刺盘孢属常规PCR检测体系。建立的PCR体系能从刺盘孢属菌株中扩增出449bp的特异性条带,刺盘孢属的近似属菌株未能扩增到条带。灵敏度试验可检测到DNA的浓度为2.2pg/0x0E?SymbolmA@0x0FL。【结论】用建立的PCR体系对炭疽病梨果实样品进行检测,可从发病组织中检测到刺盘孢属真菌。建立的PCR方法可靠、灵敏度高,能够快速、准确检测出刺盘孢属真菌。

【Objective】Colletotrichum fungi are important plant pathogens that cause plant anthracnose.This project aims to establish a PCR detection method for the Colletotrichum.【Method】The specific primers of the Colletotrichum were designed by comparing and analyzing the internal transcribed spacers(ITS)sequences of the Colletotrichum and allied genera,and the PCR amplification system was established to verify the specificity and sensitivity of the primers.【Result】A pair of species-specific primer ITS-c3/c4 was designed for Colletotrichum and a conventional PCR detection system was established.The established PCR system could amplify a specific band of 449 bp from the Colletotrichum,but the allied genera could not do so.The concentration of DNA was 2.2 pg/L by sensitivity test.The established PCR system was used to detect the anthracnose pear fruit samples and the fungi of the Colletotrichum could be detected from the pathogenic tissues.【Conclusion】The established PCR method is reliable and sensitive,which can be used to quickly and accurately detect the Colletotrichum.