雄性生殖干细胞（mGSCs）是研究不育症和精子发生的机理及转基因动物生产的理想细胞模型，而揭示其自我更新的精密调控机理是体外建立稳定雄性生殖干细胞株的前提。我们已初步发现PLZF为奶山羊mGSCs的自我更新所必需，但尚不清楚其确切的分子机制。本项目拟采用RNAi 和超表达PLZF，解析PLZF对奶山羊雄性生殖干细胞的自我更新的作用；利用PLZF的抗体进行ChIP-Seq，寻找PLZF在染色体上的特异结合和调控位点，结合RNA-Seq技术筛选PLZF自我更新相关下游靶基因；以体外培养的mGSCs和三维培养的生精小管为材料，RNAi 、过表达、荧光素酶实验等对筛选的关键靶基因进行功能研究和验证; MeDIP、ChIP测定PLZF对自我更新相关靶基因的甲基化，组蛋白甲基化和乙酰化等表观遗传修饰的作用。阐明PLZF维持奶山羊mGSCs自我更新的分子机理，为提高家畜mGSCs的建系效率提供科学依据。

Male germline stem cells (mGSCs), also known as spermatogonia stem cells (SSCs), is an ideal cell model for the study of infertility and spermatogenesis, and generation of transgenic animal research. To explore the mechanisms of self-renewal of mGSCs is the prerequisite of in vitro establishment of stable mGSC lines. Previous studies have shown that PLZF is necessary for the self-renewal of mice and dairy goat mGSCs , however, the specific molecular mechanism of which hasn’t been defined yet. Therefore, this project tries to investigate the molecular mechanism of PLZF by using RNAi and over-expression of PLZF; We investigate to use ChIP-Seq by PLZF antibody to search for the specific binding site on a chromosome and regulation of PLZF; in combination with RNA-Seq technique to sreen for the downstream target gene of PLZF; to test and verify the screening prospective candidate targeted genes of PLZF by knocking-down and over-expression them in vitro culturing of mGSCs and the three-dimension culture of seminiferous tubules, and luciferase assay; and further to determine effects of epigenetic modifications including the DNA methylation, histone methylation and acetylization on PLZF by using MeDIP and ChIP technologies; and eventually to reveal the molecular mechanism of PLZF on the regulation of the self-renewal of dairy goat mGSCs. This study will provide scientific evidences to improve the efficiency of establishment of livestock mGSC line.