草鱼全基因组微卫星特征分析与亲子鉴定  

Analysis of microsatellite in the entire grass carp(Ctenopharyngodon idella)genome and the application in parentage identification

作　　者：黄纬杰 郭向召 张子豪 董强 熊雪梅 高泽霞[1,2] HUANG Weijie;GUO Xiangzhao;ZHANG Zihao;DONG Qiang;XIONG Xuemei;GAO Zexia(Key Laboratory of Freshwater Animal Breeding,Ministry of Agriculture and Rural Affairs,Engineering Research Center of Green Development for Conventional Aquatic Biological Industry in the Yangtze River Economic Belt,Ministry of Education,Engineering Technology Research Center for Fish Breeding and Culture in Hubei Province,College of Fisheries,Huazhong Agricultural University,Wuhan 430070,China;Hubei Hongshan Lab,Wuhan 430070,China;Animal Husbandry and Fisheries Research Center of Guangdong Haid Group Co.,Ltd.,Guangzhou 511400,Chin)

机构地区：[1]华中农业大学水产学院,农业农村部淡水生物繁育重点实验室,长江经济带大宗水生生物产业绿色发展教育部工程研究中心,湖北省名优鱼育种与健康养殖工程技术研究中心,湖北武汉430070 [2]湖北洪山实验室,湖北武汉430070 [3]广东海大集团股份有限公司畜牧水产研究中心,广东广州511400

出　　处：《水产学报》2022年第2期161-172,共12页Journal of Fisheries of China

基　　金：国家自然科学基金(31872559);国家大宗淡水鱼类产业技术体系建设项目(CARS-46-08);中国博士后科学基金(2019M652829)。

摘　　要：为分析草鱼全基因组微卫星特征并开发高多态性微卫星标记亲子鉴定平台,实验利用已发布的草鱼全基因组序列,开发高度多态、准确度高、重复单元在4~6碱基范围的微卫星标记。结果显示,在草鱼900.51 Mb基因组序列中共筛选到微卫星序列677363个,总长度12835407 bp,占全基因组长度的1.4254%,平均跨度为1329.43 bp。其中单碱基重复序列出现最多,占比52.85%;二碱基重复序列其次,占比31.44%;再次是四碱基重复序列,占比8.05%,三碱基重复序列占比6.47%,五碱基重复序列占比1.12%;六碱基重复序列出现最少,仅占比0.07%。1~6碱基重复类型中优势重复单位分别是A/T、AC/GT、AAT/ATT、AGAT/ATCT、AATAT/ATATT、AACCCT/AGGGTT。随机挑选重复序列为4~6个碱基的110个位点设计引物,在草鱼繁殖亲本群体中扩增,筛选出15对多态性引物,经CERVUS 3.0软件分析15个位点平均期望杂合度0.802~0.959,模拟分析结果鉴定准确率为100%,置信度为95%。通过对20个家系子代192个个体进行亲本来源鉴定,所有子代均成功匹配到正确父母本,鉴定准确率达到100%。本研究分析了草鱼全基因组微卫星特征,并利用多态性微卫星位点进行草鱼亲子鉴定,为草鱼微卫星的功能和应用研究以及草鱼的育种工作奠定基础。Grass carp(Ctenopharyngodon idella)is the most productive freshwater aquaculture species in China,which is of great importance for Chinese aquaculture industry.Developing molecular marker assisted breeding program for C.idella has important theoretical value and practical significance.Microsatellite markers are popular in breeding studies with characterization of wide distribution,large numbers and high polymorphism.In this study,microsatellite loci were mined based on whole genome sequences of C.idella and the distribution characteristics of the loci were analyzed.In the meantime,microsatellite markers with 4-6 bases repeats with high polymorphism and high accuracy of paternity test were developed.The results showed that 677363 microsatellite sequences were found in the 900.51 Mb genome sequence of C.idella,with a total length of 12835407 bp,accounting for 1.4254%of the whole genome length.The average distance of SSR was 1329.43 bp.The mononucleotide repeats motifs(52.85%)were the most common,followed by dinucleotide repeat(31.44%),tetranucleotide repeat(8.05%),trinucleotide repeat(6.47%),pentanucleotide repeat(1.12%),hexanucleotide repeat(0.07%).The preponderance of 1-6 base repeat motifs were A/T,AC/GT,AAT/ATT,AGAT/ATCT,AATAT/ATATT and AACCCT/AGGGTT,respectively.The distributions of copy numbers in different microsatellites repeat types showed there was a significant negative correlation between repeats motifs length and mean copy number(r=-0.8581,P=0.0288).The copy numbers of mononucleotide repeats were focused mainly on 10-29(98.11%),dinucleotide repeats 6-25(96.80%),trinucleotide repeats 5-11(95.14%),tetranucleotide repeats 5-10(88.48%),pentanucleotide repeats 5-10(84.93%),hexanucleotide repeats 5-7(87.42%).The 110 loci with 4-6 base repeats were randomly selected and primers were designed.Among these loci,50 loci were amplified successfully with mixed DNA of C.idella used as a template and 15 loci showed high polymorphsim when analyzed in its breeding parent population.Cervus 3.0 analysis results showed th

关 键 词：草鱼 全基因组 微卫星 亲子鉴定 

分 类 号：Q785[生物学—分子生物学] S917.4[农业科学—水产科学]