樱桃SRAP-PCR体系优化及其遗传多样性分析

选取亲缘关系较远的3个不同基因型樱桃资源为试材,对影响SRAP标记PCR反应的模板、Mg2+、dNTPs、Taq酶及引物浓度进行了优化,建立了适合于樱桃SRAP标记的扩增体系。反应体系具体为:模板DNA75ng,dNTPs0.2mmol·L-1,Mg2+2.5mmol·L-1,引物0.3μmol·L-1,Taq酶1.0U,反应总体积20μL。采用优化的扩增体系,对45个樱桃种质材料进行了遗传多样性分析,筛选8对扩增清晰且多态性高的引物组合,检测位点共227个,其中多态性位点192个,占84.6%。应用NTSYS-pc软件进行聚类分析(UPGMA),结果表明45个樱桃品种可分为欧洲甜樱桃和中国樱桃2大类,品种间遗传相似系数在0.52～0.98;其中中国樱桃与甜樱桃种间的相似系数最小,表明2类种质具有不同的遗传背景;而组群内的不同品种资源表现了较高的遗传相似性。SRAP分子标记的聚类分析揭示了樱桃品种间亲缘关系与地理分布以及来源相关。

Optimization of SRAP-PCR system and its application in genetic diversity analysis of cherry

Three different genotypes of cherry germplasm showing distant relationship were used as material for studying the effects of concentrations of Mg2+,dNTPs,Taq DNA polymerase,primers and DNA template on the SRAP-PCR reactions and optimizing the establishment of SRAP molecular marker system in cherry.The optimum system was established as follows:template DNA 75 ng,dNTPs 0.2 mmol·L-1,Mg2+ 2.5 mmol·L-1,primer 0.3 μmol·L-1,Taq polymerase 1.0 U,the total reac-tion volume was 20 μL.Amplifications were carried out on 45 samples with eight primer combinations;using this optimum system.227 steady and reliable bands were amplified,of which 192(84.6%) were polymorphic.UPGMA cluster analysis was performed by soft NTYSIS-pc 2.01,the clades formed including two groups at the dice coefficient of 0.52,namely sweet cherry(Prunus avium L.) and Chinese cherry(P.pseudocerasus L.).The genetic similarity coefficient of the 45 accessions ranged from 0.52 to 0.98.The lowest genetic similarity coefficient existed between sweet cherry and Chinese cherry;it showed that the genetic background of two groups differed from each other.The high similarity coefficient existed among dif-ferent cultivars belonging to the same groups.The cluster results revealed by SRPA showed that the relationship of cherry germplasm was related to geographic distribution and originals.