基于定制高密度SNP芯片的绒山羊绒用性状主效基因的鉴定及功能分析

Array-based genome-wide association study is powerful approach in the genetic analyses of livestock species. The currently available 53K SNP chip for goats (Illumina) is a medium density array (SNP=53,347), and all the SNP information were collected from the goat breeds outside of China, leading to the difficulties of genetic studies with respect to the Chinese goat breeds. Based on our goat resequencing studies using eight breeds, this project aims to choose around 80,000 SNPs out of 7,383,653 SNPs which were yielded by resequencing of Inner Monglia cashmere goats. These selected SNPs will subject to build a custom array for cashmere goats, termed Cashmerearray. Genome-wide association study will be subsequently performed using the Cashmerearray in a highly established cashmere goat population (n=1000) with cashmere traits recorded, in order to identify the both positional and functional candidate genes underlying cashmere traits including cashmere yield, diameter and length of cashmere. Furthermore, the function of candidate major genes will be validated by laboratory techniques in molecular and/or cellar levels. Our results will benefit the application of molecular breeding in cashmere goats by improving cashmere yield and the quality of cashmere, as well as provide valuable information in transgenic goat studies via gene editing approach.

基于SNP芯片的全基因组关联分析（GWAS）目前是动物基因遗传分析的有效工具。现有的商用山羊芯片因SNP密度较低（53K）且SNP信息均源于国外山羊品种，用于开展中国山羊品种的遗传分析存在明显缺陷，因此国内尚未开展相关研究。本项目以业已获得的8个山羊品种的重测序数据为基础，从其中内蒙古白绒山羊738万多个SNP中选择约8万个具有潜在功能的SNP进行绒山羊高密度芯片的制备。再利用该芯片对一个具有6年完整产绒量、羊绒细度和羊绒长度等绒用性状记录的绒山羊群体（n=1000）进行GWAS分析，旨在发现影响绒山羊绒用性状的（位置型和功能型）主效基因。同时，利用分子生物学和细胞生物学方法对主效基因进行验证和功能分析。本项目的实施有助于在常规育种基础上开展绒山羊的分子育种，快速提高绒山羊产绒量和羊绒品质。同时，可为利用基因编辑技术进行转基因绒山羊育种奠定基础。

在本项目支持下，筛选获得了绒山羊特有的可用于芯片制作的中国绒山羊SNP集合，该集合共包括176,200个SNPs，这些SNPs平均分布在山羊的基因组中，可用于绒山羊的基因组选择及全基因组关联分析，并已通过合作方式（与内蒙古农业大学和北京畜牧兽医研究所）用于生产中国山羊SNP芯片。通过对近1000只具有连续5-6年产羔记录的内蒙古白绒山羊选择出连续5-6年产羔数大于2的个体15只，选择连续5-6年产羔数为1的个体15只，进行基因组重测序，获得了一批与绒山羊高繁殖力性状高度相关的选择基因。同时，通过分析全球家养山羊、野生山羊物种和野生山羊历史遗迹的基因组，为一个类似高加索羱羊物种到家养山羊祖先的一个古老渗入痕迹提供了证据。在前期基础上，通过对绒山羊毛囊细胞（毛乳头细胞、毛囊干细胞和毛母质细胞）体外培养体系更进一步完善，并可用于绒山羊毛囊发育相关功能的验证。我们还做了进一步拓展研究，利用已经鉴定的与绒山羊经济性状相关的功能基因，采用CRISPR、单碱基编辑等基因编辑方法对相关基因进行定向编辑。