桑树EST—SSR引物开发

【目的】利用桑科的EST数据库进行桑树EST—SSR引物开发，为丰富桑树SSR数据库提供参考。【方法】下载NCBI中桑科的EST数据库，进行序列处理及拼接后，用SSRIT搜索其中的SSR位点，以Primer Premier 5．0设计引物，选用5个桑树品种各5株样品进行多态性引物筛选。【结果】下载的EST拼接后得到2008条拼接片段（contig），共搜索到831个SSR位点，分布在799个contig中。二、三核苷酸重复基元是主导类型，分别占总SSR的66．06％和32．37％；TC／AG和CT／GA是二核苷酸优势重复基元，分别占二核苷酸总数的28．64％和25．87％。设计合成77对EST—SSR引物，其中46对能够扩增出有效条带；经多态性筛选，26对引物可扩增出多态性条带。【结论】利用桑属近缘属的EST序列进行桑树EST—SSR引物的开发是可行的；开发出的26对多态性引物可用于桑树遗传多样性分析和种质鉴定等。

Development of EST-SSR primers in mulberry

Objective]In this study, EST-SSR primers of mulberry were developed based on the public Moraceae EST data to provide references on enriching SSR marker data in this species. Method ]ESTs of Moraceae were downloaded from NCBI database. After being revised and rejoined, SSR locations were searched using SSRIT and primers were designed using Primer Premier 5.0 software. Polymorphic primers were selected using 25 materials of five varieties. Result] Being revised and rejoined, 2008 non-redundant contigs were gained. Based on those contigs, a total of 831 SSR loci distributed in 799 contigs were detected. Dinucleotide and trinucleotide repeats were the dominant types among the obtained EST-SSRs, accounting for 66.06% and 32.37%, respectively. TC/AG and CT/GA were the most abundant in dinucleotide, reaching to 28.64% and 25.87%, respectively. Seventy-seven primer pairs were designed and synthesized, among which 46 pairs could amplify effective bands. After polymorphic selection, 26 pairs primers were polymorphic. Conclusion]It was an effective and feasible approach to develop SSR markers based on ESTs in Moraceae. Twenty-six polymorphic primer pairs were developed, which could be used in analysis on genetic diversity, germplasm identification, etc.