The capacity of a single-celled zygote to divide, specify, and generate an organism is a remarkable, yet intricate, process. While the mouse embryo has shed light on many questions of mammalian embryogenesis, human early cell fate specification and morphogenesis remain a mystery, given the legal and ethical concerns of human embryo research. During the last forty years, pluripotent stem cells have become an invaluable tool to mimic early cell fate specification in a petri dish and to depict the molecular mechanisms of early development. Furthermore, in the last decade, combining 3D matrix scaffolds with specific cell types and origins has created diverse 2D and 3D stem cell-based models that effectively mimic key aspects of mammalian embryogenesis.

In this Methods Collection, we cover protocols for the direct generation of early mouse and human embryonic and extraembryonic cell types that arise during the preimplantation and early post-implantation stages. This approach includes recent advances in the generation of integrated and non-integrated stem cell-based embryo models, as well as approaches to generate specific cell types in 2D by pharmacological, morphogenetic, and genetic modulation of key signaling cascades. We aim to provide a clear and practical roadmap for generating early mammalian lineages and developing robust stem cell-based models that capture defined developmental windows.