Extended Data Fig. 1: Development of culture conditions for chicken ESC derivation.

a, Representative images of chicken blastodermal cells cultured in the indicated conditions for 3 days. Scale bars, 50 μm. b, A representative image of chicken blastodermal cells cultured in IWR-1 + Gö6983 for 3 days. The right panel provides a zoom-in view of the dotted area within the image on the left panel. Scale bars, 100 μm (left panel), 50 μm (right panel). c, Detection of Nanog mRNA expression by RNAscope in situ hybridization in chicken blastodermal cells cultured in the indicated conditions for 3 days. Scale bars, 100 μm. d, e, Representative images of chicken blastodermal cells cultured in the indicated conditions for 10 days, with passaging once (d), or without passaging (e). Scale bars, 50 μm. f, A representative image of chicken blastodermal cells cultured in IWR + Gö6983 supplemented with 3% yolk (Yolk/2i) collected from a fertile egg (passage 1, total 10 days). Scale bar, 50 μm. g, RT-qPCR analysis of Nanog mRNA expression in chicken ESCs cultured with or without Yolk/2i for 5 days. EGK.X and E5 embryos were included as positive and negative controls, respectively. Data represent mean ± s.d. from three biological replicates. Cell images in a-f are from one representative experiment of at least 3 experiments performed.