奶山羊全基因组拷贝数变异（CNVs）挖掘及泌乳相关基因鉴定

Copy number variations, a form of structural variation are alterations of the DNA that results in the cell having an abnormal number of copies of one or more sections of the DNA, compared to SNP, they have more important genetics effect. Many studies research on copy number variation and their genetic effects, they find that copy number variations are not only association with human and animal diseases but also play important roles in phenotypic characters. The goat (Capra hircus) represents one of the most important farm animal species. It is reared in all continents with an estimated world population of about 800 million of animals. Shannxi province have the most number of dairy goat in our country, and its milk products play important role in economy. But nowadays, the phenomenon of variety degeneration and production performance decreased deeply effect industry development. Many experts research on it, but because short of genome sequence, studies of the goat genome are still in their infancy compared to those in other farm animal species. In December 2012, the first goat genome has been sequenced and the data has been released. These will cause research focus on goat genome. Using goat genomic data, we want to detect copy number variations in different breeds of dairy goat (Guanzhong dairy goat, Laoshan dairy goat, Saanen dairy goat) by array-based comparative genomic hybridization (aCGH) combined with bioinformatics methods. The chips will compose with three breeds, each breed has three individuals, and the results will validate by QPCR. Structure and distribution characteristics of CNVs (numbers of copy, CNV across more than one gene, CNV located in coding, regulatory or intergenic region) will be analyzed in our program and we will describe a first map of goat CNVs. Using Gene Ontology (GO) to analyze CNVs which located in gene coding or regulatory region to find economic related pathway's gene. 15-20 lactation relevant genes' CNV will choose to detect its polymorphism in goat population with production records. And their association with economic traits will be analyzed in more than six hundreds goat population in this study. We will choose CNVs which showed significant results to analyze its genetic mechanism. Based on different CNVs locations and structures (copy numbers, CNV located in coding or regulatory regions), we will use bioinformatics and molecular biology methods to reveal the molecular mechanism. This study will help to protect germplasm resource and will provide new ideas for molecular breeding in goat, especially the important lactation related genes which validate in our study will offer application value to dairy goat. Our results have important academic and application prospect.

基因组拷贝数变异（Copy number variation,CNV）对人和动物疾病的发生及许多性状的表现具有重要影响。2012年12月底，世界上首个山羊全基因组测序完成且序列已释放，这将引起山羊基因组数据挖掘的研究热点。本项目拟依据山羊全基因组序列，采用比较基因组杂交芯片与生物信息学相结合的方法发现奶山羊基因组中CNVs，分析其结构特点及分布特征，QPCR方法进行验证，绘制山羊基因组CNVs图谱。GO富集分析得到与泌乳性能相关信号通路中基因的CNVs；挑选15-20个泌乳通路中基因的CNVs，利用MLPA技术及统计软件分析实验群体中奶山羊泌乳性能与CNVs的关联性。对具有显著效应的CNVs，根据CNVs在染色体及基因中的位置设计实验进行分析，阐明CNVs 的遗传机制，筛选泌乳相关基因进行鉴定，本研究将为山羊种质资源保护和奶山羊分子育种及品种改良提供遗传基础，结果具有重要的学术及应用价值。

2012年，世界上首个山羊全基因组测序完成且序列已释放，这将引起山羊基因组数据挖掘的研究热点。本研究基于2012年发布的山羊全基因组序列，通过山羊全基因组测序结合生物信息学、分子生物学的方法分析并检测山羊全基因组单核苷酸多态性（SNPs）、插入缺失（indels）和拷贝数变异（CNVs） 的数目、群体特征、结构特征及分布规律，并挖掘影响山羊泌乳性能的重要CNVs 标记及关键基因。研究将开创中国山羊基因组变异的新领域，并对奶山羊重要产奶性状进行遗传解析。.通过测序分析了15只家养山羊（5只辽宁白绒山羊、5只萨能奶山羊、5只雷州肉山羊）及4只野山羊个体（巴基斯坦野山羊）。每个个体30乘的数据，达到90G左右的数据量，总数据量达1800G的数据。通过BWA 0.5.9，ANGSD，Dindel v1.01，Control-FREEC等多个生物信息学软件进行比对及SNPs, Indels, CNVs分析：共检测到23万多的SNP位点变异；检测到1-5bp缺失插入变异2400多万个；依据大于1kb核酸片段重复为CNV，共发现2137个拷贝数变异位点。对于群体间差异的CNV片段进行验证，发现位于13号染色体上一个CNV片段包含ASIP和AHCY基因，这两个基因均与毛色相关，进一步进行验证发现，白色羊毛山羊品种（萨能奶山羊，辽宁绒山羊）在这个位点的拷贝数目明显高于黑色山羊品种（雷州山羊，巴基斯坦DDT品种）。对于奶山羊特有的CNV多态性进行群体的关联分析，发现APOL3 和NME6基因拷贝数变异与关中奶山羊产奶量和生长显著显著相关。将家养山羊15个样本与4个巴基斯坦野山羊群体进行比较，利用固定指数及群体杂合度进行统计计算，发现67个驯化相关基因，这些基因涉及神经发育，免疫和形态等多个方面。萨能奶山羊与其他两个品种相比，发现Fst>0.5的区段16个，涉及26个基因，KEGG通路富集后，亚油酸代谢通路极显著。辽宁绒山羊与其他两个品种相比，发现Fst>0.5的区段10个，涉及14个基因。雷州山羊与其他两个品种相比，发现Fst>0.5的区段16个，涉及27多个基因，KEGG通路富集后，甘油磷酸代谢通路显著。以上发现的候选基因均可进行深入的功能研究，本项目将为山羊种质资源保护和奶山羊分子育种及品种改良提供遗传基础，结果具有重要的学术及应用价值。