Generation of human PSCs in EPSCM. (A) Representative images of H1-FGF (left panel) or H1-EPSCM (right panel) PSC colonies (scale bar = 50 μm). (B) Cell cycle analysis of H1-FGF and H1-EPSCM (passage 15 in EPSCM) PSCs. Data representative of three biological replicates. (C) Graphical representation of the reprogramming strategy using episomal or piggyBac transposon vectors for PSC reprogramming from neural stem cells (NSCs) and human dermal fibroblasts (HDFs), respectively. Primary colonies appeared after 10–15 days, at which time the media was switched to EPSCM for an additional 6–8 days before picking. (D) Images of a primary PSC colony in EPSCM (left panel) and established EC5-EPSCM PSCs (right panel) (scale bar = 100 μm). (E, F) Expression of pluripotency and lineage markers genes in H1-FGF, H1-EPSCM, EC5-EPSCM, and PB-EPSCM PSCs. Parental HDFs and NSCs were used as control. Relative expression of these genes was normalized to GAPDH. Data are mean ± standard deviation. Data are representative of three biological replicates. (G) Immunostaining of H1-EPSCM PSCs for the cell surface markers SSEA3, SSEA4, and Tra-181 and the pluripotency transcription factors OCT4, SOX2 and NANOG (scale bar = 200 μm). (H) Spectral karyotype of H1-EPSCM and EC5-EPSCM PSCs at passage 12 and passage 15, respectively.