scRNA-seq reveals transcriptional dynamics of Notch-stimulated LPs. (A) UMAP visualization of scRNA-seq data from Cas9-LPs with or without Notch stimulation (left) and thymus-derived primary DN cells (Kit^high: Kit^low = 1:2) (right). Clustering was performed using a nonlinear representation of the top 50 principal components. Cells are colored according to the time after Notch stimulation (left). (B) Pseudo-time scores of individual cells are shown in UMAP1-2, with colors indicating progression along pseudo-time. Pseudo-time scores were calculated using Monocle3, with the principal root node defined as LPs without Notch stimulation. (C) Distribution of Cas9-LPs, Notch-stimulated T progenitors, and primary thymic DN cells along pseudo-time (top). The relative expression patterns of Spi1, Bcl11b, and Tcf7 across pseudo-time are shown (bottom). (D) Heatmap showing SCENIC-predicted regulon activities of the indicated TFs across different developmental stages of DN cells. Data are based on two independent experiments, and pooled data were used for analysis.