Extended Data Fig. 2: Validation of 4CL and e4CL media.
From: Rolling back human pluripotent stem cells to an eight-cell embryo-like stage
a. Heatmap showing the expression of preimplantation ICM-enriched genes in human naïve PSC cultured in NHSM29, tt2iLGö (this study), 5iLAF17 and 4CL (passage 5), and human ICM cells16 compared with primed PSC. All reference datasets used in this study are summarized in Supplementary Table 8. H9 ESC were used to generate our dataset. The full list of DEG is included in Supplementary Table 1. b. RT-qPCR validation of totipotency genes in primed H9 ESC cultured in stepwise e4CL (day 5). Data are presented as mean values ± SD of fold-change compared to 4CL-day 12 naïve ESC. n = 3 biological replicates. c. Heatmap showing the expression of totipotency genes in naïve ESC cultured in NHSM, tt2iLGö, 5iLAF or stepwise e4CL (day 5), EPSC11, sorted 8CLC from e4CL-day 5 cells, and human 8C-embryo cells16 compared with primed PSC. H9 ESC were used to generate our dataset. d. Representative images of G-banding karyotype of primed H9 ESC and iPSC-4 cultured in stepwise e4CL (day 5). Twenty metaphases were counted for each. All 20 metaphase spreads for H9 ESC are shown in Supplementary Figure 1. e. RT-qPCR validation of totipotency genes in primed H9 ESC cultured in direct e4CL (day 7). Data are presented as mean values ± SD of fold-change compared to primed ESC. n = 3 biological replicates.
