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为了从分子水平探讨苹果炭疽病病原菌的群体遗传多样性,采用L16(45)正交设计对ISSR-PCR反应体系中的Mg2+浓度、dNTP浓度、Taq DNA聚合酶用量、引物浓度和DNA含量进行优化,并利用优化的体系进行引物筛选及苹果炭疽病菌遗传多样性分析。确立最优反应体系为2.0mmol/L Mg2+、0.2 mmol/L dNTP、2 U Taq DNA聚合酶、1μmol/L引物和DNA 100 ng。筛选获得的10条引物对供试菌株共扩增出42条谱带,均为多态性条带。供试菌株在相似系数0.50处分为2个类群,分别与根据形态学鉴定的胶孢刺盘孢Colletotrichum gloeosporioides和尖孢刺盘孢C.acuta-tum 2个类群相一致,在相似系数0.74处,分为4个亚群,表明苹果炭疽病菌存在明显的种间及种内遗传分化。
In order to explore the population genetic diversity of apple anthracnose pathogens at the molecular level, the L16 (45) orthogonal design was used to optimize the Mg2 + concentration, dNTP concentration, Taq DNA polymerase, primer concentration and DNA content in the ISSR-PCR reaction system , And optimized the system for primer selection and analysis of genetic diversity of apple anthracnose. The optimal reaction system was 2.0 mmol / L Mg2 +, 0.2 mmol / L dNTP, 2 U Taq DNA polymerase, 1 μmol / L primer and 100 ng of DNA. A total of 42 bands were amplified by the 10 primer pairs screened, which were all polymorphic bands. The tested strains were divided into two groups at a similarity coefficient of 0.50, which were consistent with the two groups of Colletotrichum gloeosporioides and C. acuta-tum identified by morphology, respectively. At the similarity coefficient of 0.74, Divided into four subgroups, indicating that apple anthracnose obvious interspecific and intraspecific genetic differentiation.