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To understand the most recent advances in germline differentiation from human pluripotent stem-cell lines in vitro, it is necessary to give a brief introduction of the basics of germline development in mammals. In mammals, germ cells arise from a pluripotent cell population called primordial germ cells (PGCs) that segregates from the somatic lineage during gastrulation in response to the Bone Morphogenetic Proteins 4 and 8 secreted by adjacent extra-embryonic endoderm. Once germ cells colonize gonadal ridges, expression of SRY, encoded on the short arm of the Y chromosome, drives their male sexual differentiation. Meiosis in mammals is a complicated process consisting in two consecutive cell divisions without DNA replication to generate haploid cells. This chapter reviews the main advances in germline differentiation from pluripotent stem cells, with special emphasis on the challenges to overcome in order to achieve the correct maturation of germ cells in vitro.