Extended Data Fig. 9. CellOracle validation using experimentally measured cell density in Tal1 KO in Pijuan-Sala et al. gastrulation and organogenesis scRNA-seq data.

(a) UMAP plot of chimeric E8.5 embryos of wild-type (WT) and Tal1 KO cells (25,307 cells and 26,311 cells, respectively) from a published scRNA-seq atlas of mouse gastrulation and organogenesis³⁰. (b) Tal1 gene expression (log-transformed UMI) projected onto the UMAP plot. (c) Pseudotime gradient vector field used in the perturbation score (PS) calculations. Developmental pseudotime was calculated using the DPT method with WT chimera scRNA-seq data and then converted into a 2D gradient vector field. (d) PS and cell transition vector field of the Tal1 KO simulation. (e) The magnified area of erythrocyte differentiation predicts inhibition or arrest of cell differentiation at the haematoendothelial progenitor stage. (f) The Markov random walk simulation result predicts high cell density in the haematoendothelial progenitor cluster and lower cell density at later stages, indicating that Tal1 KO would induce differentiation arrest at the haematoendothelial progenitor stage. (g) Experimentally measured Tal1 KO data. The kernel cell density of whole chimera (left), WT (middle), and Tal1 KO cells (right) were calculated after downsampling each condition (25,307 cells) to control for sample size. A scatter plot of whole chimera cells is shown as background (light grey) to highlight the overall cell trajectory structure. (h) The bar plot shows the cell type composition in each sample (right panel). Overall, the experimental result aligns with the simulated predictions. The relative fold change between WT and KO samples is also shown in Supplementary Table 4. (i) Perturbation score and cell transition vector field of the Tal1 conditional KO simulation in the erythroid lineage. Tal1 expression was set to zero in the Blood progenitor and Erythrocyte clusters; CellOracle simulates KO effects in later erythroid differentiation stages. (j) The Markov simulation result shows uniform cell density, predicting that Tal1 KO would not induce differentiation arrest in a conditional KO targeting later stages of erythroid differentiation.