Fig. 1: Intensive scRNA-seq of somite-resolved E8.5 mouse embryos.

a, A new scRNA-seq dataset was generated from nuclei derived from individual E8.5 mouse embryos via an optimized sci-RNA-seq3 protocol to bridge existing data generated on E8.5 cells via 10x Genomics² and E9.5 nuclei via sci-RNA-seq3 (ref. ⁴). b, 3D UMAP visualizations of the new E8.5 dataset (E8.5b). All nuclei colored by germ layer are shown in the center, along with separate embeddings of neuroectoderm (left), nonhematopoietic mesoderm (bottom right) and endoderm, extraembryonic and hematopoietic cell types (top right). c, Twelve mouse embryos, including a single primitive-streak-stage embryo and 11 embryos staged in 1-somite increments from 2 to 12 somites, were collected and their nuclei subjected to optimized sci-RNA-seq3. d, Re-embedded two-dimensional (2D) UMAP of cells annotated as forebrain, midbrain, hindbrain, spinal cord and neural crest. Arrows correspond to RNA velocity trends⁹⁷. e, The same UMAP as in d, colored by somite counts. The subset of cells from rhombomere 4 that appear to emerge the earliest are highlighted in red circles (Hoxa1⁺ and Hoxb1⁺)^23,24. f, For each cell type with >100 profiled cells, we calculated the Pearson correlation coefficient between the somite number of each cell of that type and the average somite number of its five nearest neighbors in the global 3D UMAP embedding. Colors indicate germ layers. g, 3D visualization of the top three PCs of gene expression variation in NMPs, calculated on the basis of the 2,500 most highly variable genes. Cells are colored by the somite count of the originating embryo. Genes most strongly correlated (Pearson), either positively (red) or negatively (green), with each PC are listed. ExE, extraembryonic; r2–r5: rhombomeres 2–5.