全基因组重测序筛选山羊经济性状候选基因

发布时间：2018-05-14 11:17

本文选题：山羊 + 基因组重测序　；参考：《西南大学》2017年硕士论文

【摘要】：我国有悠久的山羊驯化和养殖历史,拥有丰富多彩的品种和遗传资源。山羊为人类提供肉、纤维、皮和奶等产品。然而目前对山羊特征体型体貌、重要的生产和繁殖等经济性状的遗传机理研究多以单基因和候选基因验证为主,这严重忽略了生物的系统性,所以筛选的致因基因的准确度也较低,从而导致山羊经济性状遗传剖分和遗传机理研究进展缓慢。随着高通量测序技术的不断发展,为从全基因组层面筛选经济性状候选基因提供了可能。本研究以饲养在西南大学实验羊场的大足黑山羊(n=6)和内蒙古绒山羊(n=6)为研究对象,采集颈静脉全血样本用于DNA提取,经检测合格后用于构建350bp双末端测序文库,然后采用Illumina HiSeq PE150平台进行全基因组测序。原始数据通过BWA软件进行质检和过滤接头等后比对到参考基因组。接着利用SAMtools软件检测基因组单核苷酸多态性(Single Nucletide Polymorphism,SNP)并用ANNOVAR软件注释。分别采用Cluster 3.0、ADMIXTURE和MEGA 5.2进行主成分分析、遗传结构和邻近进化树分析;变异比较分析首先筛选每个群体相同的SNPs,然后比较两个群体差异SNPs,并将外显子SNPs注释到基因;其次分别计算群体杂合性(Heterozygosity,Hp)和群体分化(Fst)来筛选候选基因,再分别用Goseq(Bioconductor 2.12)和KOBAS(kobas2.0 20120208)进行GO和KEGG注释分析初步了解候选基因功能。本研究共产生192.747Gb原始数据,平均鉴定了500多万个SNPs。主成分分析、遗传结构和邻近进化树分析均显示大足黑山羊和内蒙古绒山羊能够被完全分成结构清晰的两个不同群体;变异比较分析发现大足黑山羊和内蒙古绒山羊特有的SNPs各有1,873和1,706个,其中外显子变异各有21和17个,分别注释到16和14个基因;杂合性计算在大足黑山羊和内蒙古绒山羊基因组分别筛选到155和294个候选区域,分别包含86和97个候选基因;Fst统计检测到368个选择消除区域,含有候选基因164个;然后选取低杂合度和高遗传分化的前1.0%区域作为受选择区域,在大足黑山羊和内蒙古绒山羊基因组内分别筛选到239和176个候选区域,分别注释到135和176个候选基因。经过GO和KEGG功能富集、并结合参考文献报道和试验验证分析发现候选基因与山羊的体貌、生产和繁殖性能相关。如KIT基因(rs647214940)可作为山羊白毛的候选基因,FGF5基因是调控山羊被毛生长的关键候选基因;BIRC6基因可能是山羊季节性繁殖的重要调控基因;EGLN1基因(rs659097694)是内蒙古绒山羊适应性高海拔环境的候选基因。本研究通过全基因重测序、变异检测、群体结构、变异比较和选择信号分析等手段,筛选了系列与山羊经济表型性状相关的候选基因,为深度揭示山羊表型性状的遗传机理奠定了基础。
[Abstract]:China has a long history of domestication and breeding of goats, with rich varieties and genetic resources. Goats provide human beings with products such as meat, fiber, skin and milk. However, at present, the genetic mechanism of economic traits such as characteristic body shape, important production and reproduction of goats is mainly verified by single gene and candidate gene, which neglects the systematicness of living things. Therefore, the accuracy of the selected genes is also low, which leads to slow progress in the research of genetic division and genetic mechanism of economic traits in goats. With the development of high throughput sequencing technology, it is possible to screen candidate genes for economic traits from the whole genome level. In this study, Dazu Black Goat (Dazu Black Goat 6) and Inner Mongolia Cashmere Goat (Cashmere Goat No. 6), which were raised in the Experimental Sheep Farm of Southwest University, were studied. The whole blood samples of jugular vein were collected for DNA extraction, and the 350bp double-terminal sequencing library was constructed after the detection. Then the whole genome was sequenced using Illumina HiSeq PE150 platform. The raw data were checked and filtered by BWA software and then compared to the reference genome. SAMtools software was used to detect single Nucletide polymorphism (SNPs) and ANNOVAR software was used to annotate SNPs. Cluster 3.0 and MEGA 5.2 were used for principal component analysis, genetic structure and adjacent phylogenetic tree analysis, variation comparison analysis was used to screen the same SNPs in each population, then the differences between two populations were compared, and the exon SNPs was annotated into the gene. Secondly, population heterozygosity (HP) and population differentiation (FST) were calculated respectively to screen candidate genes, and then go and KEGG annotation analysis were carried out with Goseq(Bioconductor 2.12) and KOBAS(kobas2.0 2010 20208) respectively to understand the function of candidate genes. In this study, the original 192.747Gb data were generated, and more than 5 million SNPs were identified on average. Principal component analysis, genetic structure and adjacent evolutionary tree analysis showed that Dazu Black Goat and Inner Mongolia Cashmere Goat could be divided into two distinct populations. The SNPs of Dazu Black Goat and Inner Mongolia Cashmere Goat were 1873 and 1706 respectively, of which exon variation was 21 and 17 respectively, annotated 16 and 14 genes, respectively. The heterozygosity was estimated to be 155 and 294 candidate regions in the genome of Dazu Black Goat and Inner Mongolia Cashmere Goat, respectively. Fst analysis of 86 and 97 candidate genes detected 368 candidate elimination regions and 164 candidate genes in the genome of Dazu Black Goat and Inner Mongolia Cashmere Goat. Then the first 1.0% region with low heterozygosity and high genetic differentiation was selected as the selected region. 239 and 176 candidate regions were screened in the genome of Dazu Black Goat and Inner Mongolia Cashmere Goat respectively, and 135 and 176 candidate genes were annotated respectively. Through the enrichment of go and KEGG functions, combined with reference reports and experimental verification analysis, it was found that candidate genes were related to goat body appearance, production and reproductive performance. For example, KIT gene rs647214940 can be used as candidate gene for goat white hair, FGF5 gene is the key candidate gene for controlling goat hair growth, and BIRC6 gene may be an important regulating gene for seasonal reproduction of goat. Candidate genes for high altitude environments. In this study, a series of candidate genes related to economic phenotypic traits of goats were screened by means of whole gene resequencing, mutation detection, population structure, variation comparison and selection signal analysis. It lays a foundation for revealing the genetic mechanism of goat phenotypic traits in depth.
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S827

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