Extended Data Figure 3: Gene expression dynamics during HSC formation.

a, Heat map of expression levels of the FACS markers for ten-cell-pool RNA-seq analysis. b, Heat map of expression levels of FACS markers for single-cell RNA-seq analysis. c, Unsupervised hierarchical clustering of transcriptome profiles of 35 ten-cell samples showing that the samples were accurately grouped together according to their cell types. d, Unsupervised hierarchical clustering of transcriptome profiles of 108 single-cell samples comprising ECs, T1 pre-HSCs, T2 CD41^low cells, E12 HSCs, and E14 HSCs. e, Bar plot of RNA expression of Pecam1 (CD31), Itga2b (CD41), and Ptprc (CD45) in the two subpopulations of T2 CD41^low cells. f, Endothelial or HSC and HPC markers specifically expressed in CD201-positive T2 CD41^low cells as well as other HSC-competent populations, and granulocyte and macrophage markers specifically expressed in the CD201-negative T2 CD41^low subpopulation. g, Donor-derived chimaerism in peripheral blood of recipients in the three-cell T2 pre-HSC group (CD31⁺CD45⁺c-Kit⁺CD201^high) monitored at 4, 8, 12, and 16 weeks after transplantation. h, Quantification of T2 pre-HSC frequency by limiting dilution analysis.

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