大菱鲆微卫星标记的分离及其多态性位点检测  

Isolation of novel microsatellite markers from genome library and SSR polymorphic loci identification in turbot Scophthalmus maximus

作　　者：顾颖[1,2] 郭忠宝[2] 谷晶晶[2] 毛瑞鑫[1,3] 鲁翠云[1] 孙效文[1] 

机构地区：[1]黑龙江水产研究所农业部北方鱼类生物工程育种重点开放实验室,黑龙江哈尔滨150070 [2]大连水产学院生命科学与技术学院,辽宁大连116023 [3]上海海洋大学生命科学与技术学院,上海201306

出　　处：《大连水产学院学报》2009年第4期366-370,共5页Journal of Dalian Fisheries University

基　　金：国家科技条件平台建设项目(2006DKA30470-005)

摘　　要：以生物素标记的（CA）12为探针进行杂交，借助磁珠筛选出含微卫星的Mbo I酶切片段，构建了大菱鲆Scophthalmus maximus微卫星富集文库，用同位素γ-^23P标记的探针（CA）12进行二次筛选，获得1600个阳性克隆，占59．3％。测序347个序列，得到335个（96．54％）含有微卫星序列的克隆，其中含有微卫星座位378个。完美型的微卫星序列有236个（62．4％），非完美型的有121个（32．0％），复合型的有21个（5．6％），从所得序列中分离和鉴定了32对微卫星标记。利用31尾大菱鲆养殖个体评价微卫星位点，分析表明，10个位点具有多态性。不同位点等位基因数目为3～11不等，期望杂合度（He）和多态信息含量（PIC）变动范围分别为0．5061—0．8995和0．4133～0．8742。本研究中开发的微卫星多态性较高，将为大菱鲆养殖品系的优化、遗传多样性的检测及遗传图谱的构建等打下基础。We constructed the microsatellites - enriched libraries that contained Mbo I - digested microsatellites sequences identified by the approach of streptavid in magnesphere paramagnetic particles and bloted with the oligo- probes of biotinylated （CA）12 in turbot （Scophthalmus maximus L. ）. Total of 1600 positive colonies （59.3%） were made through twice screens by radioactive γ-^ 32 P- ATP labeling hybridization. Sequencing of 347 positive colonies confirmed that there were 335 colonies containing 378 microsatellite loci in which 21 were compound repeat motifs （5.6%） , 236 were perfect （62.4%） and 121 were imperfect （32.0%）, respectively. Thirty- two pairs of PCR primers were designed by software package, and microsatellite variation was assessed in 31 individuals of reared turbot strains, and 10 microsatellite loci were turned out to be polymorphic. The number of alleles per polymorphic locus ranged from 3 to 11, and the values of He and PIC varied from 0.5061 to 0. 8995 and 0.4133 to 0. 8742 , respectively. The polymorphic microsatellites isolated may contribute to future studies of population genetic structure and the construction of the genetic mapping for the turbot.

关 键 词：大菱鲆 微卫星 多态性 磁珠 

分 类 号：S917[农业科学—水产科学]