双须骨舌鱼转录组EST-SSR标记开发与引物筛选  

Development and primer selection of EST-SSR molecular markers based on transcriptome sequencing of Osteoglossum bicirrhosum

作　　者：王且鲁[1,2] 刘奕[1] 宋红梅[1] 汪学杰[1] 刘超[1] 牟希东[1] 胡隐昌[1] 罗建仁[1] 

机构地区：[1]农业部热带亚热带水产资源利用与养殖重点实验室,中国水产科学研究院珠江水产研究所,广州510380 [2]上海海洋大学水产与生命学院,上海201306

出　　处：《淡水渔业》2016年第6期8-13,共6页Freshwater Fisheries

基　　金：广东省自然科学基金(2014A030310397);国家水产种质资源平台(2016DKA30470)

摘　　要：利用MISA软件对双须骨舌鱼(Osteoglossum bicirrhosum)性腺转录组测序获得的188 461条Unigene进行SSR检测,结果在11 917个Unigene中共检测到SSR位点12 578个,其中包含2个及以上SSR位点的Unigene有633个。SSR的发生频率为6.32%,平均分布距离为9 949 bp。在筛选到的SSR位点中,优势重复基序为二核苷酸、三核苷酸和四核苷酸,其中二核苷酸重复基序数量最多,占总数的50.52%。重复基元类型共216种,其中数量最多的类型为二核苷酸重复AC/GT,占总数的20.66%。从所有筛选出的SSR位点中随机选取50个位点并运用Primer Premier 5.0设计引物,共有25对引物成功扩增出特异性产物,并且均在同科的美丽硬仆骨舌鱼(Scleropages formosus)的三个亚种中通用。结果表明,转录组测序产生的EST序列是开发SSR标记的有效来源,开发所得EST-SSR引物可用于双须骨舌鱼及其近缘物种遗传分析、遗传图谱构建以及分子标记辅助育种等工作中。A total of 188 461 unigenes from gonad transcriptome of Osteoglossum bicirrhosum were screened using MISA software. 12 578 EST-SST molecular markers were identified in 11 917 Unigenes,and the frequency of these SSRs was6. 32% and mean distance was 0. 99 kb in the unigenes. Six hundred and thirty three unigenes contained more than one SSR. Dinucleotide,trinucleotide and tetranucleotide repeats were dominant motifs,and the most frequent repeat was dinucleotide which accounted for 50. 52% of the total number of SSRs. A total of 216 types of motifs were detected and AC /GT was presented to be the most frequent motif,accounting for 20. 66%. A total of 50 SSRs were selected for design primers by Primer Premier 5. 0,and 25 pairs of which were effective,which could be transferable to three subspecies of Scleropages formosus. This study indicated that EST sequences produced by transcriptome sequencing was an effective source to develop SSR markers. The 25 SSR markers developed in this study can be used in genetic analysis,linkage map construction and marker assisted breeding of O. bicirrhosum and its closely related species.

关 键 词：双须骨舌鱼(Osteoglossum bicirrhosum) 转录组 EST-SSR 

分 类 号：S917.0[农业科学—水产科学]