West Jason A, Park In-Hyun, Daley George Q, Geijsen Niels
Graduate Program of Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts, USA.
Nat Protoc. 2006;1(4):2026-36. doi: 10.1038/nprot.2006.303.
The demonstration of germ cell and haploid gamete development from embryonic stem cells (ESCs) in vitro has engendered a unique set of possibilities for the study of germ cell development and the associated epigenetic phenomenon. The process of embryoid body (EB) differentiation, like teratoma formation, signifies a spontaneous differentiation of ESCs into cells of all three germ layers, and it is from these differentiating aggregates of cells that putative primordial germ cells (PGCs) and more mature gametes can be identified and isolated. The differentiation system presented here requires the differentiation of murine ESCs into EBs and the subsequent isolation of PGCs as well as haploid male gametes from EBs at various stages of differentiation. It serves as a platform for studying the poorly understood process of germ cell allocation, imprint erasure and gamete formation, with 4-6 weeks being required to isolate PGCs as well as haploid cells.
体外从胚胎干细胞（ESC）中证明生殖细胞和单倍体配子的发育为生殖细胞发育及相关表观遗传现象的研究带来了一系列独特的可能性。胚状体（EB）分化过程，如同畸胎瘤形成一样，意味着ESC自发分化为所有三个胚层的细胞，正是从这些正在分化的细胞聚集体中，可以识别和分离出假定的原始生殖细胞（PGC）和更成熟的配子。这里介绍的分化系统要求将小鼠ESC分化为EB，并随后从不同分化阶段的EB中分离出PGC以及单倍体雄配子。它作为一个平台，用于研究人们了解较少的生殖细胞分配、印记消除和配子形成过程，分离PGC和单倍体细胞需要4至6周时间。
