大豆疫霉基因组SSR标记开发

Development of Genomic SSR Markers for Phytophthora sojae

【目的】开发大豆疫霉基因组SSR标记,为从分子水平深入研究大豆疫霉及其近缘种提供一种理想的分子标记。【方法】用FPCR软件从大豆疫霉全基因组序列中查询SSRs,选择合适的SSR序列用Primer5.0软件设计引物。【结果】从发现的1234个含有2～4个碱基重复单元的完全SSRs中选出260段设计引物,经10个大豆疫霉分离物基因组DNA检测,有213对(81.9%)扩增出SSR特征条带,其中114(53.5%)对引物扩增出多态性。通用性检测表明,14.6%～28.6%引物分别在选择的8个疫霉种中有效扩增。基于10个SSR标记数据进行聚类分析,结果表明表明这些标记可以完全区分大豆疫霉及其它疫霉。【结论】大豆疫霉基因组SSR标记具有高多态性,是大豆疫霉遗传变异、遗传多样性、及遗传图谱构建等研究理想工具。部分大豆疫霉基因组SSR标记在其近缘种中具有通用性,可以用于疫霉菌的系统进化、鉴定、区分及开发近源种的SSR标记。特别开发的SSR标记,在大豆疫霉基因组中具有准确的位置,将极大地方便大豆疫霉菌功能基因的定位和克隆,以及进行疫霉菌的比较基因组的研究。

【Objective】 This study was aimed at developing genomic SSR markers for Phytophthora sojae and offering ideal molecular markers for studying it and relative species in depth on molecular level. 【Method】 Using Fast PCR software to screening SSRs in complete genome sequence of P. sojae, suitable sequence regions harboring SSR were selected for design of markers using Primer5.0 software, and synthesized primers were used to amplify genomic DNA of selected isolates of P. sojae and other Phytophthora species. [ Result ] Of the designed 260 primer pairs, 213 （81.9%） could amplify characteristic SSR fragments against 10 different isolates ofP. sojae, and 114 （53.5%） primer pairs amplified polymorphic fragments among the 10 isolates. The results for transferability test showed that 14.6%-28.6% primer pairs could effectively amplify in 8 selected Phytophthora species, respectively. Clustering Phytophthora spp and P. sojae based on 10 SSR markers data, these markers could clearly differentiate selected Phytophthora species. [Conclusion] The genomic SSR markers of P. sojae are highly polymorphic, and should be the ideal tools for studies of genetic variation, genetic diversity, and development of genetic linkage maps of the pathogen. Some SSR primers have transferability in other Phytophthora species, and could be used in phylogenetic analysis, species identification and differentiation and SSR marker development of Phytophthora. Especially, these genomic SSR markers have been located in P. sojae genome, thus facilitating lagging and cloning the functional genes of P. sojae, and carrying comparative genomics study of Phytophthora.