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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。
Siniperca chuatsi is an important freshwater cultured fish species in China. In order to identify the pedigree genealogies of Culter alburnus, the identification results of double digest restriction-site associated DNA sequencing (ddRADseq) and microsatellite markers were compared, In this study, 11 pairs of primers with specificity and polymorphism were designed and screened based on the microsatellite sequences in GenBank, and amplified, typed and verified in 4 sib families with known pedigree information. The clustering results showed that the 11 pairs of microsatellite primers could divide the four full-sib families into four branches, which is in accordance with the actual pedigree. A total of 3 283 SNP markers were obtained using ddRAD-seq to construct a cluster tree containing 155 sibling individuals and 12 other non-sibling individuals, which can be effectively distinguished. The above 11 pairs of microsatellite primers were used to detect the 12 non-sibling individuals, 8 sibling individuals and their parents, and the clustering tree was constructed. The results showed that 8 sibling individuals were clustered with their parents and 12 non-sibling individuals were clustered For the other one, indicating that the 11 microsatellite loci have better pedigree identification ability. The 11 pairs of microsatellite primers were used for paternity testing of 126 offspring breed by 7 male and 13 female breeding parents. The simulated results showed that the rate of non-adherence of parents in each microsatellite locus was 0.276 ~ 0.895, The highest cumulative rate of non-kinship of parents was 99.999%. The results of paternity test showed that 105 out of 126 progeny successfully constructed pedigree information, and the success rate of identification was 83.3%, indicating that these 11 sites not only had higher cumulative exclusion rate , In practical applications can also have a better identification results. In this study, the establishment of parental appraisal technology of Culter alburnus will provide effective pedigree identification technology for Breeding Culter alburnus.