鲑鳟通用型低通量单核苷酸多态性芯片的开发
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摘要
为开发常见鲑鳟养殖物种通用的遗传分析工具,本研究利用Affymetrix虹鳟(Oncorhynchus mykiss) 57K高通量单核苷酸多态性(Single nucleotide polymorphism,SNP)芯片,对国内代表性鲑鳟养殖群体开展了分型检测,包括山女鳟(Oncorhynchus masou masou)、银鲑(Oncorhynchus kisutch)、美洲红点鲑(Salvelinus fontinalis)、白斑红点鲑(Salvelinus leucomaenis) 4个物种,从57,501个SNP标记中筛选出96个共享多态性标记,应用Fluigidm 96.96动态芯片平台,构建了大麻哈鱼属(Oncorhynchus)和红点鲑属(Salvelinus)通用型低通量SNP芯片。该芯片分型结果准确性较高,与Affymetrix高通量芯片分型一致性达到96.55%。使用该芯片对来自6个家系的48尾银鲑个体及其候选亲本进行检测,应用Cervus 3.0.7软件进行亲权鉴定,结果能够准确重现复杂家系的真实系谱。在用于单亲本亲权鉴定时,第一亲本非排除率(Non-exclusion probability for first parent, NE-1P)为4.120×10~(–4);用于双亲本亲权鉴定时,双亲非排除率(Non-exclusion probability for parent pair, NE-PP)低至6.219×10~(–12),表明该芯片在鲑鳟养殖群体系谱鉴定应用中具有较高的准确性。使用该芯片开展4个鲑鳟养殖群体遗传结构分析,样本分群聚类结果与其所属的分类阶元相符,能够准确反映群体遗传组分构成和遗传关系。本研究构建的低通量SNP芯片在常见鲑鳟养殖物种中具有良好的通用性,将其应用于养殖群体遗传分析,能够为鲑鳟制种、育种和引种等科学决策提供基因组信息参考。
This study aimed to develop a universal genotyping tool for common salmonid species cultured in China. Using an Affymetrix rainbow trout(Oncorhynchus mykiss) 57 K single nucleotide polymorphism(SNP) array, four representative aquaculture salmonid populations were genotyped, including Oncorhynchus masou masou, Oncorhynchus kisutch, Salvelinus fontinalis, and Salvelinus leucomaenis. A total of 96 shared polymorphism markers was screened out of 57,501 SNPs, then a low-density SNP array was constructed based on a Fluigidm 96.96 dynamic array, which is universal for the genera Oncorhynchus and Salvelinus. Reliability of the genotyping results was high because the identity of results with those of the Affymetrix 57 K array reached 96.55%. Parentage assignment for 48 Oncorhynchus kisutch offspring was performed with Cervus 3.0.7 software, based on genotyping data by the 96 SNP array. The results matched perfectly with the true pedigrees among complicated families, proving high accuracy in the pedigree assignment application of the 96 SNP array. The non-exclusion probability for the first parent(NE-1P) was 4.120×10~(–4) during single parent assignment, whereas the non-exclusion probability for the parent pair(NE-PP) was as low as 6.219×10~(–12). Genetic structure analysis for four salmonid populations was also performed using genotyping data of the 96 SNP array. Clustering results were in accordance with the taxonomic categories of the samples, whereas certain degrees of genetic introgression were observed among the populations, indicating that the genetic composition and relationships among populations were properly demonstrated in the analysis. All these results suggested that the developed 96 SNP array has great potential in genetic analyses for salmonid populations, which might provide helpful genome-wide information for plasm resource evaluation, selective breeding, high-quality fry production, and reasonable foreign strain introduction in the Chinese salmonid aquaculture.
This study aimed to develop a universal genotyping tool for common salmonid species cultured in China. Using an Affymetrix rainbow trout(Oncorhynchus mykiss) 57 K single nucleotide polymorphism(SNP) array, four representative aquaculture salmonid populations were genotyped, including Oncorhynchus masou masou, Oncorhynchus kisutch, Salvelinus fontinalis, and Salvelinus leucomaenis. A total of 96 shared polymorphism markers was screened out of 57,501 SNPs, then a low-density SNP array was constructed based on a Fluigidm 96.96 dynamic array, which is universal for the genera Oncorhynchus and Salvelinus. Reliability of the genotyping results was high because the identity of results with those of the Affymetrix 57 K array reached 96.55%. Parentage assignment for 48 Oncorhynchus kisutch offspring was performed with Cervus 3.0.7 software, based on genotyping data by the 96 SNP array. The results matched perfectly with the true pedigrees among complicated families, proving high accuracy in the pedigree assignment application of the 96 SNP array. The non-exclusion probability for the first parent(NE-1P) was 4.120×10~(–4) during single parent assignment, whereas the non-exclusion probability for the parent pair(NE-PP) was as low as 6.219×10~(–12). Genetic structure analysis for four salmonid populations was also performed using genotyping data of the 96 SNP array. Clustering results were in accordance with the taxonomic categories of the samples, whereas certain degrees of genetic introgression were observed among the populations, indicating that the genetic composition and relationships among populations were properly demonstrated in the analysis. All these results suggested that the developed 96 SNP array has great potential in genetic analyses for salmonid populations, which might provide helpful genome-wide information for plasm resource evaluation, selective breeding, high-quality fry production, and reasonable foreign strain introduction in the Chinese salmonid aquaculture.
引文
Falush D,Stephens M,Pritchard JK.Inference of population structure using multi locus genotype data:Linked loci and correlated allele frequencies.Genetics,2003,164(4):1567-1587
Harlizius B,Lopes MS,Duijvesteijn N,et al.A single nucleotide polymorphism set for paternal identification to reduce the costs of trait recording in commercial pig breeding.Journal of Animal Science,2011,89(6):1661-1668
Hu G,Gu W,Bai QL,et al.Research advances in genetics and breeding in salmonids.Chinese Journal of Fisheries,2012,25(3):58-62[户国,谷伟,白庆利,等.主要养殖鲑科鱼类遗传育种的研究进展.水产学杂志,2012,25(3):58-62]
Hubisz MJ,Falush D,Stephens M,et al.Inferring weak population structure with the assistance of sample group information.Molecular Ecology Resources,2009,9(5):1322-1332
Kalinowski ST,Taper ML,Marshall TC.Revising how the computer program CERVUS accommodates genotyping error increases success in paternity assignment.Molecular Ecology,2007,16(5):1099-1106
Li DY,Kong J,Meng XH,et al.Development of multiplex PCRsystems of microsatellite markers for Pacific white shrimp(Litopenaeus vannamei)and its application for parentage identification.Progress in Fishery Sciences,2016,37(3):58-67[李东宇,孔杰,孟宪红,等.凡纳滨对虾(Litopenaeus vannamei)微卫星多重PCR体系的建立及其在家系亲权鉴定中的应用.渔业科学进展,2016,37(3):58-67]
Liang LQ,Chang YM,Dong CZ.Analysis of genetic diversity for Coregonus usssruensis Berg in Heilongjiang River.Journal of Fishery Sciences of China,2004,11(6):501-505[梁利群,常玉梅,董崇智.黑龙江乌苏里白鲑遗传多样性分析.中国水产科学,2004,11(6):501-505]
Liu S,Palti Y,Gao G,et al.Development and validation of a SNP panel for parentage assignment in rainbow trout.Aquaculture,2016,452:178-182
Lu CY,Cao DC,Li YF,et al.Evaluation of genetic potential of Siberian char Salvelinus leucomaenis cultured population using microsatellite markers from rainbow trout Oncorhynchus mykiss.Chinese Journal of Fisheries,2016,29(1):1-7[鲁翠云,曹顶臣,李永发,等.利用虹鳟微卫星标记评价白斑红点鲑养殖群体的遗传结构.水产学杂志,2016,29(1):1-7]
Naish KA,Phillips RB,Brieuc MS,et al.Comparative genome mapping between Chinook salmon(Oncorhynchus tshawytscha)and rainbow trout(O.mykiss)based on homologous microsatellite loci.G3(Bethesda).2013,3(12):2281-2288
Palti Y,Gao G,Liu S,et al.The development and characterrization of a 57K single nucleotide polymorphism array for rainbow trout.Molecular Ecology Resources,2015,15(3):662-672
Purcell S,Neale B,Todd-Brown K,et al.PLINK:A tool set for whole-genome association and population-based linkage analyses.American Journal of Human Genetics,2007,81(3):559-575
Si F,Sun ZH,Yu SS,et al.Analysis of the genetic variability of the captured population of Japanese flounder Paralichthys olivaceus in the coastal releasing area of Qinhuangdao.Progress in Fishery Sciences,2017,38(6):18-24[司飞,孙朝徽,于姗姗,等.秦皇岛近海增殖放流区牙鲆(Paralichthys olivaceus)回捕群体的遗传多样性分析.渔业科学进展,2017,38(6):18-24]
Small MP,Beacham TD,Withler RE,et al.Discriminating coho salmon(Oncorhynchus kisutch)populations within the Fraser River,British Columbia,using microsatellite DNAmarkers.Molecular Ecology,2010,7(2):141-155
Sun DJ,Wang BQ.Aquaculture of salmonids in China.Chinese Journal of Fisheries,2010,23(2):56-63[孙大江,王炳谦.鲑科鱼类及其养殖状况.水产学杂志,2010,23(2):56-63]
Wang J,Wang QY,Kong J,et al.SSR analysis on genetic diversity in breeding and wild populations of Fenneropenaeus chinensis.Progress in Fishery Sciences,2017,38(6):18-24[王军,王清印,孔杰,等.中国对虾人工选育群体与野生群体遗传多样性的SSR分析.渔业科学进展,2018,39(2):1-8]
Xu J,Feng J,Peng W,et al.Development and evaluation of a high-throughput single nucleotide polymorphism multiplex assay for assigning pedigrees in common carp.Aquaculture Research,2017,48(4):1866-1876
Xu J,Zhao Z,Zhang X,et al.Development and evaluation of the first high-throughput SNP array for common carp(Cyprinus carpio).BMC Genomics,2014,15(1):307
Yu JN,Azuma N,Yoon M,et al.Genetic structure and phylogeography of masu salmon(Oncorhynchus Masou masou)inferred from mitochondrial and microsatellite DNAAnalyses.Zoological Science,2010,27(5):375-385
Harlizius B,Lopes MS,Duijvesteijn N,et al.A single nucleotide polymorphism set for paternal identification to reduce the costs of trait recording in commercial pig breeding.Journal of Animal Science,2011,89(6):1661-1668
Hu G,Gu W,Bai QL,et al.Research advances in genetics and breeding in salmonids.Chinese Journal of Fisheries,2012,25(3):58-62[户国,谷伟,白庆利,等.主要养殖鲑科鱼类遗传育种的研究进展.水产学杂志,2012,25(3):58-62]
Hubisz MJ,Falush D,Stephens M,et al.Inferring weak population structure with the assistance of sample group information.Molecular Ecology Resources,2009,9(5):1322-1332
Kalinowski ST,Taper ML,Marshall TC.Revising how the computer program CERVUS accommodates genotyping error increases success in paternity assignment.Molecular Ecology,2007,16(5):1099-1106
Li DY,Kong J,Meng XH,et al.Development of multiplex PCRsystems of microsatellite markers for Pacific white shrimp(Litopenaeus vannamei)and its application for parentage identification.Progress in Fishery Sciences,2016,37(3):58-67[李东宇,孔杰,孟宪红,等.凡纳滨对虾(Litopenaeus vannamei)微卫星多重PCR体系的建立及其在家系亲权鉴定中的应用.渔业科学进展,2016,37(3):58-67]
Liang LQ,Chang YM,Dong CZ.Analysis of genetic diversity for Coregonus usssruensis Berg in Heilongjiang River.Journal of Fishery Sciences of China,2004,11(6):501-505[梁利群,常玉梅,董崇智.黑龙江乌苏里白鲑遗传多样性分析.中国水产科学,2004,11(6):501-505]
Liu S,Palti Y,Gao G,et al.Development and validation of a SNP panel for parentage assignment in rainbow trout.Aquaculture,2016,452:178-182
Lu CY,Cao DC,Li YF,et al.Evaluation of genetic potential of Siberian char Salvelinus leucomaenis cultured population using microsatellite markers from rainbow trout Oncorhynchus mykiss.Chinese Journal of Fisheries,2016,29(1):1-7[鲁翠云,曹顶臣,李永发,等.利用虹鳟微卫星标记评价白斑红点鲑养殖群体的遗传结构.水产学杂志,2016,29(1):1-7]
Naish KA,Phillips RB,Brieuc MS,et al.Comparative genome mapping between Chinook salmon(Oncorhynchus tshawytscha)and rainbow trout(O.mykiss)based on homologous microsatellite loci.G3(Bethesda).2013,3(12):2281-2288
Palti Y,Gao G,Liu S,et al.The development and characterrization of a 57K single nucleotide polymorphism array for rainbow trout.Molecular Ecology Resources,2015,15(3):662-672
Purcell S,Neale B,Todd-Brown K,et al.PLINK:A tool set for whole-genome association and population-based linkage analyses.American Journal of Human Genetics,2007,81(3):559-575
Si F,Sun ZH,Yu SS,et al.Analysis of the genetic variability of the captured population of Japanese flounder Paralichthys olivaceus in the coastal releasing area of Qinhuangdao.Progress in Fishery Sciences,2017,38(6):18-24[司飞,孙朝徽,于姗姗,等.秦皇岛近海增殖放流区牙鲆(Paralichthys olivaceus)回捕群体的遗传多样性分析.渔业科学进展,2017,38(6):18-24]
Small MP,Beacham TD,Withler RE,et al.Discriminating coho salmon(Oncorhynchus kisutch)populations within the Fraser River,British Columbia,using microsatellite DNAmarkers.Molecular Ecology,2010,7(2):141-155
Sun DJ,Wang BQ.Aquaculture of salmonids in China.Chinese Journal of Fisheries,2010,23(2):56-63[孙大江,王炳谦.鲑科鱼类及其养殖状况.水产学杂志,2010,23(2):56-63]
Wang J,Wang QY,Kong J,et al.SSR analysis on genetic diversity in breeding and wild populations of Fenneropenaeus chinensis.Progress in Fishery Sciences,2017,38(6):18-24[王军,王清印,孔杰,等.中国对虾人工选育群体与野生群体遗传多样性的SSR分析.渔业科学进展,2018,39(2):1-8]
Xu J,Feng J,Peng W,et al.Development and evaluation of a high-throughput single nucleotide polymorphism multiplex assay for assigning pedigrees in common carp.Aquaculture Research,2017,48(4):1866-1876
Xu J,Zhao Z,Zhang X,et al.Development and evaluation of the first high-throughput SNP array for common carp(Cyprinus carpio).BMC Genomics,2014,15(1):307
Yu JN,Azuma N,Yoon M,et al.Genetic structure and phylogeography of masu salmon(Oncorhynchus Masou masou)inferred from mitochondrial and microsatellite DNAAnalyses.Zoological Science,2010,27(5):375-385
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