Abstract
An in vitro cell model mimicking human 8C blastomeres would be invaluable to understanding the mechanisms regulating major zygote genome activation (ZGA) in humans. Here, we present a protocol for deriving human preimplantation epiblast stem cells and 8-cell embryo-like cells. We describe steps for developing an 8C-specific GFP reporter (8C::GFP) and capturing the rare human 8C-like cells (8CLCs) from human preimplantation epiblast-like stem cells (prEpiSCs). We then detail procedures for optimizing the culture condition to extend 8CLC culturing. For complete details on the use and execution of this protocol, please refer to Yu et al.1.