鸡原始生殖细胞向精子诱导分化研究

In vitro induced sperm differentiated from stem cell lines possesses huge values for preparing transgenic animals and treating male infertility. At present, the molecular regulatory elements and the regulation pathways of spermatogenesis have not been cleared, the culture system in vitro is still hard to reach the requirement of the natural developing environment, which left two challenges, "functional differentiation" and "differentiation efficiency". We have successfully obtained induced sperm in vitro through retinoic acid from chicken primordial germ cells (PGCs), which provided the foundation for this research. We aim to establish a new sperm induction technology which including several steps presented below: The key genes and the signal paths during spermatogenesis would be screened out via the genetic sequencing technology and the molecular mechanism would be explored, then recombinant vectors contained those genes would be constructed for transfering into PGCs and the in vitro testis organ culture method would be combined, thus, this kind of technical route might solve the problems above, which has a great significance for improving avian genetics and breeding and transgenic varieties.

干细胞体外诱导分化为精子在转基因动物制备、雄性不育治疗上具有重大的应用价值。由于对精子发生的分子调控元件及调控途径不祥，体外培养体系难以达到体内环境要求，使目前的研究尚未在"功能成熟"和"分化效率"两个难点上取得突破。我们已成功分离纯化了鸡原始生殖细胞(Primordial germ cell,PGC)通过维甲酸诱导获得了具有运动能力精子。本项目将以此为研究基础，紧密围绕以上2个关键科学问题，首次通过转录组测序技术，筛选精子发生过程中的关键因子和信号通路，并探索精子发生的分子机制；在此基础上采用转基因策略将构建的关键因子重组载体导入PGC中，建立单（或多）因子控制PGC向精子分化的诱导技术。对转染的PGC探索利用体外器官培养法诱导其精子发生的技术体系，以促进诱导精子的"功能成熟"。本项目对于推进家禽遗传育种、品种改良以及转基因品种培育具有重要意义。

干细胞体外诱导分化为精子在转基因动物制备、雄性不育治疗上具有重大的应用价值。但目前体外定向分化为精子的研究，仍存在“功能成熟”和“分化效率”两个难点。本项目紧密围绕这2个拟解决的关键科学问题，对鸡精子发生关键调控基因进行系统研究，建立体外鸡的原始生殖细胞（PGCs）定向分化为功能性生殖细胞，从而推进家禽遗传育种、品种改良的发展。自获得国家自然金基金资助以来，该项目实施进展顺利，成功建立了PGCs体外扩增的方法，建立体外诱导PGCs向精子发生技术，并通过单细胞测序，获得原始生殖细胞向雄性生殖细胞诱导分化的功能因子。