Extended Data Fig. 3: Transcriptomic analysis of the transcriptional features of S2 and S3 cells and the effect of compound from S2-S3 base.

a, Pseudotime trajectory reconstruction of single cells from subpopulations at S1 and S2. b, Box plots showing the expression of genes enriched in subpopulation 1 at S1 (S1-1) across subpopulations from S1 and S2. Each box depicts the median level along with the first and third quartiles. The whiskers represent the minimum and maximum values. Points indicate outliers, which are defined as values exceeding 1.5 times the interquartile range from the box. The dots in the boxplot represent genes. p value was calculated using the two-sided Wilcoxon rank-sum test, n = 100 (number of genes). p values from left to right: 2.96 × 10⁻¹⁷, 3.04 × 10⁻²², 2.16 × 10⁻²⁷, 7.28 × 10⁻²⁴. c, Heatmaps showing the expression of totipotency marker genes in S2 cell aggregates under different chemical combinations. S2, CHIR-99021+Birabresib+CD1530 + 8-Br-cAMP; -C, Birabresib+CD1530 + 8-Br-cAMP; -B, CHIR-99021 + CD1530 + 8-Br-cAMP; -CD, CHIR-99021+Birabresib+8-Br-cAMP; -8Br, CHIR-99021+Birabresib+CD1530. d, Heatmaps showing the expression of Nr5a2 and Tfap2c target genes in S2 cell aggregates under different chemical combinations. S2, CHIR-99021+Birabresib+CD1530 + 8-Br-cAMP; -C, Birabresib+CD1530 + 8-Br-cAMP; -B, CHIR-99021 + CD1530 + 8-Br-cAMP; -CD, CHIR-99021+Birabresib+8-Br-cAMP; -8Br, CHIR-99021+Birabresib+CD1530. e, Expression of marker genes for epiblast, trophectoderm and primitive endoderm in S3 cell aggregates under different chemical combinations. S3, CHIR-99021+Birabresib+CD1530 + 8-Br-cAMP; -C, Birabresib+CD1530 + 8-Br-cAMP; -B, CHIR-99021 + CD1530 + 8-Br-cAMP; -CD, CHIR-99021+Birabresib+8-Br-cAMP; -8Br, CHIR-99021+Birabresib+CD1530.

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