| Germ cells are the origin of individual organisms and the carriers of genetic information and genetic diversity.Life continues with the fertilization between oocyte and sperm to produce new individuals.Therefore,the production and development of germ cells are crucial to the reproduction of species.Sperm and eggs are derived from primordial germ cells(PGCs)at the embryonic stage.Their development and maturation are regulated by many factors,including transcription factors,DNA methylation,and epigenetic modification.Because of the limited number of PGCs in vivo,an alternative approach to investigate embryonic germ cell development is in vitro differentiation of embryonic stem cells(ESCs).In this study,I utilized different methods to differentiate mouse ESCs into PGC-like cells(PGCLCs),and then co-cultured them with somatic cell to induce these PGCLCs to go through spermatogonial stage to enter meiosis.I applied this this differentiation system to multiple ESC lines,and successfully established a mouse germ cell differentiation platform.The ESC in vitro differentiation platform provides an useful tool to understand regulatory mechanism of germ cells development.A thorough understanding of regulatory network in the process of PGCs specialization and development lays a foundation for studying reproductive process.To systematically identify key regulators in embryonic germ cell development,we utilized this ESC differentiation platform in combination with CRISPR/dCas9 gain-of-function screening.We obtained several genes that positively influence derivation of PGCs from ESCs.These potential regulators showed high expression in PGCLCs from ESCs and PGCs development in vivo.Our independent differentiation experiments further confirmed our screening results.Taken together,our findings will eventually improve understanding regulatory mechanism of germ cell development,and also offer novel knowledge to identify causes for reproductive diseases. |
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