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【目的】建立刺葡萄(Vitis davidii Fo?x.)的ISSR分子标记体系以及分析刺葡萄种质资源的亲缘关系,为刺葡萄的保护和利用提供依据。【方法】以8份外缘葡萄品种或类型为对照,52份刺葡萄类型为材料,采用改良CTAB法提取植物基因组DNA,以基因组DNA为模板,用同一试验考察多个因素及水平的筛选方式对ISSR-PCR扩增反应体系中的Mg2+、d NTPs、引物、模板DNA的浓度及循环数进行优化,建立适用于刺葡萄的最佳ISSR-PCR反应体系;在优化的反应体系中进行多态性引物的筛选并进行分析,同时根据Jaccard遗传相似系数进行UPGMA聚类分析。【结果】采取改良CTAB法提取的刺葡萄DNA质量优于常规的CTAB法,在优化的ISSR-PCR反应体系中,从100条ISSR引物筛选出了13个具有多态性的引物,共检测到139个位点,其中多态性位点116个,多态位点百分率为83.45%。聚类结果显示,对照组8份外缘资源均在刺葡萄种群外,刺葡萄与华东葡萄的遗传距离较腺枝葡萄近;52份刺葡萄类型分为三大组群,两大江西刺葡萄组群及湖南与福建刺葡萄组群,而在湖南与福建刺葡萄种群中又分为4个群组。【结论】刺葡萄种质资源具有丰富的遗传多样性,ISSR分子标记可有效揭示刺葡萄种质资源的遗传多样性和亲缘关系,对刺葡萄种质资源的保护和利用有重要的意义。
【Objective】 ISSR molecular marker system of Vitis davidii Fo? X. Was established and the genetic relationship of germplasm resources was analyzed to provide the basis for the protection and utilization of grape. 【Method】 Eight varieties of peripheral grape varieties or types were used as controls and 52 varieties of grapes were used as materials. The genomic DNA was extracted by modified CTAB method. The genomic DNA was used as template to screen multiple factors and levels The optimized ISSR-PCR reaction system was optimized for the concentration of Mg2 +, dNTPs, primer and template DNA in ISSR-PCR reaction system and the number of cycles. The optimized reaction system was characterized by polymorphism The primers were screened and analyzed, and the UPGMA cluster analysis was performed according to the Jaccard genetic similarity coefficient. 【Result】 The results showed that the quality of DNA extracted from Crabgrass by modified CTAB was better than that of conventional CTAB. In the optimized ISSR-PCR reaction system, 13 polymorphic primers were screened from 100 ISSR primers. 139 loci, of which 116 polymorphic loci, the percentage of polymorphic loci was 83.45%. The clustering results showed that the eight peripheral resources of the control group were outside the prickly pear population, and the genetic distance between the prickly grapes and the east grapes was closer than that of the pruning grapes. The 52 types of prickly grapes were divided into three groups, Group and Hunan and Fujian thorn grape group, but in Hunan and Fujian thorn grape population is divided into four groups. 【Conclusion】 The germplasm resources of Prunus mume were rich in genetic diversity. ISSR markers could effectively reveal the genetic diversity and genetic relationship of germplasm resources of Prunus villosa. It is of great significance to the conservation and utilization of germplasm resources of Prunus villosa.