翘嘴鳜微卫星标记亲权鉴定技术的建立与应用  

Establishment and Utilization of Paternity Identification in Mandarin Fish (Siniperca chuatsi) Using Microsatellites

作　　者：曾庆凯[1,2] 孙成飞[1] 董浚键[1] 田园园[1,2] 叶星[1,2] 

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

出　　处：《农业生物技术学报》2017年第6期976-984,共9页Journal of Agricultural Biotechnology

基　　金：现代农业产业技术体系建设专项资金(CARS-49);广东省海洋渔业科技推广专项(No.A201601A06);广东省科技计划项目(No.2015A020209034);广州市科技计划项目(No.2014J4100087)

摘　　要：翘嘴鳜(Siniperca chuatsi)是中国重要的淡水优质养殖鱼类。为鉴定翘嘴鳜选育群体系谱,比较双酶切限制性酶切位点关联DNA测序(double digest restriction-site associated DNA sequencing,ddRADseq)与微卫星标记两种方法的鉴定效果,并建立翘嘴鳜亲权鉴定技术,本研究根据GenBank上的微卫星序列,设计并筛选11对具有特异性和多态性的引物,在4个已知系谱信息的全同胞家系扩增、分型与验证。聚类结果显示,这11对微卫星引物可将这4个全同胞家系分成4个支,与实际系谱情况相符。利用ddRAD-seq获得3 283个SNP标记,构建包含155个同胞个体以及12个其他非同胞个体的聚类树,这12个非同胞个体可被有效区分。采用以上11对微卫星引物检测此12个非同胞个体、8个同胞个体及其亲本并构建聚类树,结果显示,8个同胞个体样本与其亲本聚为一支,12个非同胞个体则聚为另一支,说明此11个微卫星位点具有较好的家系鉴定能力。采用这11对微卫星引物对由7尾雄性、13尾雌性选育亲本繁育的126尾子代进行亲权鉴定,模拟分析发现,各微卫星位点的双亲非亲排除率为0.276~0.895,最高累计双亲非亲排除率为99.999%;亲权鉴定结果显示,126个子代中有105尾成功构建系谱信息,鉴定成功率为83.3%,说明这11个位点不仅具有较高的累计排除率,在实际应用中也可有较好的鉴定效果。本研究中翘嘴鳜亲权鉴定技术的建立将为翘嘴鳜选育种提供有效的系谱鉴定技术手段。Mandarin fish （Siniperca chuatsi） is one of the most important cultured freshwater fish in China. In order to carry out pedigree analysis of selective mandarin fish, compare identification effects of double digest restriction-site associated DNA sequencing （ddRAD-seq） and microsatellites, and establish patemity identification in mandarin fish, 30 pairs of microsatellite primers were selected from GenBank, and 11 of them with good specificity and polymorphism were screened by polyacrylamide gel electrophoresis. Then, these 11 primers were used to amplify 4 full-sib families whose pedigree information was known. Capillary electrophoresis was used to carry out microsatellite genotyping. The result of cluster analysis showed these four full-sib families were divided into four groups using the 11 pairs of microsatellite primers. All 3 283 SNP markers were obtained by ddRAD-seq, and they were used to construct a clustering tree, including 155 samples from the same family and 12 samples from other families. Finally, the 12 samples were effectively distinguished. The above 11 pairs of microsatellite primers were used to detect the 12 samples from other families, eight samples from the same family and their parents, and clustering tree was constructed. The result showed that the 8 samples from the same family and their parents were clustered together, while the 12 samples were clustered in the other cluster. These results illustrated that these 11 pairs of microsatellite primers had good family identification ability. Then, parentage determination of 126 offsprings, which were bred by 7 male and 13 female breeding parents, were carried out by these microsatellite loci. Simulation analysis showed that the non-exclusion probability of each locus ranged from 0.276 to 0.895, and the highest calculate exclusion probability was 99.999%. Among these 126 offsprings, 105 offsprings had been successfully identified for their pedigree relationship by parentage analysis and the success rate was 83.3%. These results illustrate

关 键 词：翘嘴鳜 亲权鉴定 微卫星 单核苷酸多态性(SNP) 双酶切限制性酶切位点关联DNA测序(ddRAD-seq) 

分 类 号：S917.4[农业科学—水产科学]