High-throughput mapping of cellular differentiation hierarchies from single-cell data promises to empower systematic interrogations of vertebrate development and disease. Here, we applied single-cell RNA sequencing to >92,000 cells from zebrafish embryos during the first day of development. Using a graph-based approach, we mapped a cell state landscape that describes axis patterning, germ layer formation, and organogenesis. We tested how clonally related cells traverse this landscape by developing a transposon-based barcoding approach (“TracerSeq”) for reconstructing single-cell lineage histories. Clonally related cells were often restricted by the state landscape, including a case in which two independent lineages converge on similar fates. Cell fates remained restricted to this landscape in chordin-deficient embryos. We provide web-based resources for further analysis of the single-cell data.

Pluripotent cells in the embryo can generate all cell types, but lineage-restricted cells are generally thought to replenish adult tissues. Planarians are flatworms and regenerate from tiny body fragments, a process requiring a population of proliferating cells (neoblasts). Whether regeneration is accomplished by pluripotent cells or by the collective activity of multiple lineage-restricted cell types is unknown. Using ionizing radiation and single-cell transplantation, we identified neoblasts that can form large descendant-cell colonies in vivo. These clonogenic neoblasts (cNeoblasts) produce cells that differentiate into neuronal, intestinal, and other known post-mitotic cell types and are distributed throughout the body. Single transplanted cNeoblasts restored regeneration in lethally irradiated hosts. We conclude that broadly distributed, adult pluripotent stem cells underlie the remarkable regenerative abilities of planarians.