基于CRISPR系统的单碱基编辑技术（BE）可在不引起DNA双链断裂（DSB）的条件下引入单个碱基的替换，避免了因DSB带来的高拷贝区DNA损伤风险。本项目利用BE技术构建山羊全基因组sgRNA文库，在所有基因中通过C>T替换引入终止密码子导致翻译终止，用于筛选和鉴定绒山羊主要毛囊细胞（毛乳头细胞和毛囊干细胞）增殖相关的必需基因。随后，利用构建的文库侵染体外培养的毛囊细胞，选择经FACS筛选后表达BE3的细胞（对照组）和继续培养5代后的细胞（试验组）进行深度测序，在全基因组水平鉴定对毛囊核心细胞增殖至关重要的必需基因。最后，利用细胞生物学方法和在绒山羊个体水平对必需基因进行功能验证和阐释。本项目以绒山羊复杂的毛囊细胞系统为研究对象，旨在从全基因组水平鉴定与毛囊增殖和分化相关的功能基因，为人工调控绒毛生长、加速绒山羊种质创新奠定基础，也将为哺乳动物毛囊生物学研究提供参考。

The recently emerged base editing (BE) technology is able to induce C-to-T conversion without double strand DNA breaks, which are toxic DNA lesions that can cause genomic rearrangements and translocations. In this project, we are going to establish a CRIPSR-STOP library comprising all genes on goat genome, in order to screen essential genes for key cell types in cashmere goat hair follicles (HF) by changing single nucleotides to create stop codons. To do this, the established library will be used to infect BE3 expressed HF cells (hair follicle stem cells and dermal papilla cells) which were derived from skin of cashmere goats in vitro. Cells infected with the CRIPSR-STOP library will be screened with FACS first (control group), then we continue to culture the cells for five passages as the treated group. Cells in both groups will be collected for deep sequencing, to identify genes that are essential for cell proliferation in the HFs of cashmere goats. Subsequently, the function of identified essential genes will be validated by gene knockout in distinct cells using molecular techniques. We are going to produce edited goats with disrupted essential genes in their HFs, and further evaluate the phenotypic and genetic differences among edited goats and control groups. The results obtained in this project will provide molecular evidence for cell proliferation and differentiation in skin HFs, and will facilitate the application of genetic improvement in cashmere goats by genome editing, as well as provide new insights into HF biology studies in mammals.