Hematopoietic stem cells (HSCs) replenish all types of blood cells. It is debating whether HSCs in adults solely originate from the aorta-gonad-mesonephros (AGM) region, more specifically, the dorsal aorta, during embryogenesis. Here we report that somite hematopoiesis, a previously unwitnessed hematopoiesis, can generate definitive HSCs (dHSCs) in zebrafish. By transgenic lineage tracing, we found that a subset of cells within the forming somites emigrate ventromedially and mix with lateral plate mesoderm-derived primitive hematopoietic cells before the blood circulation starts. These somite-derived hematopoietic precursors and stem cells (sHPSCs) subsequently enter the circulation and colonize the kidney of larvae and adults. RNA-seq analysis reveals that sHPSCs express hematopoietic genes with sustained expression of many muscle/skeletal genes. Embryonic sHPSCs transplanted into wild-type embryos expand during growth and survive for life time with differentiation into various hematopoietic lineages, indicating self-renewal and multipotency features. Therefore, the embryonic origin of dHSCs in adults is not restricted to the AGM.