文章摘要
草鱼全基因组微卫星特征分析与亲子鉴定
The analysis of microsatellite in grass carp (Ctenopharyngodon idellus) genome and the application in parentage identification
投稿时间:2020-11-17  修订日期:2021-04-01
DOI:
中文关键词: 草鱼  全基因组  微卫星  亲子鉴定
英文关键词: Ctenopharyngodon idellus  genome  microsatellite  parentage identification
基金项目:国家自然科学(31872559);国家大宗淡水鱼类产业技术体系建设项目(CARS-46-08);武汉市科技计划项目(2020020601012253);中国博士后科学(2019M652829)
作者单位邮编
黄纬杰 华中农业大学水产学院/农业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/农业动物遗传育种与繁殖教育部重点实验室 430070
郭向召 广东海大集团股份有限公司畜牧水产研究中心 
张子豪 华中农业大学水产学院/农业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/农业动物遗传育种与繁殖教育部重点实验室 
董强 华中农业大学水产学院/农业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/农业动物遗传育种与繁殖教育部重点实验室 
熊雪梅 华中农业大学水产学院/农业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/农业动物遗传育种与繁殖教育部重点实验室 
高泽霞 华中农业大学水产学院/农业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/农业动物遗传育种与繁殖教育部重点实验室
湖北省名优鱼育种与健康养殖工程技术研究中心
广东海大集团股份有限公司畜牧水产研究中心 
430070
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中文摘要:
      草鱼是我国重要的淡水养殖品种,开展分子标记辅助育种研究具有重要的理论和实践意义。本研究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900.51Mb基因组序列中共筛选到微卫星序列677 363个,总长度12 835 407 bp,占全基因组长度的1.425 4%,平均跨度为1 329.43bp。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,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。
英文摘要:
      Grass carp (Ctenopharyngodon idellus) 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. idellus has important theoretical value and practical significance. Microsatellite markers are popular in breeding studies with characterized of wide distribution, large numbers and high polymorphism. In this study, microsatellite loci were mined based on whole genome sequences of C. idellus 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 677, 363 microsatellite sequences are found in the 900.51 Mb genome sequence of C. idellus, with a total length of 12, 835, 407 bp, accounting for 1.425 4% of the whole genome length. The average distance of SSR is 1, 329.43 bp. The mononucleotide repeats motifs (52.85%) are the most common, following 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 are A/T, AC/GT, AAT/ATT, AGAT/ATCT, AATAT/ATATT and AACCCT/AGGGTT, respectively. 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. idellus used as template and 15 loci showed high polymorphsim when analyzed in its breeding parent population. Cervus 3.0 analysis results showed that average expected heterozygosity of 15 loci was 0.802 ~ 0.959. The accuracy of simulated paternity test was 100% with 95% confidence degree. These 15 loci were used to identify 192 offspring from 20 families, and all offspring matched to the correct parents successfully with the accuracy of identification being 100%. In this study, we analyzed the microsatellite characteristics of C. idellus, screened out highly polymorphic microsatellite markers with 4-6 base repeats and achieving parental identification of C. idellus by using these highly polymorphic microsatellite loci. At the same time, it was proved that 4-6-base repeat microsatellite is more suitable for paternity test than traditional 2-3-base repeat. This study lays a foundation for the application of C. idellus microsatellites in its breeding program.
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