The relative contribution of yolk sac (YS)-derived cells to the circulating definitive hematopoietic progenitor cell (HPC) pool that seeds the fetal liver remains controversial due to the presence of systemic circulation and the onset of hematopoiesis within the embryo proper (EP) before liver seeding. Ncx1-/- embryos fail to initiate a heartbeat on embryonic day (E) 8.25, but continue to develop through E10. We detected normal numbers of primitive erythroid progenitors in Ncx1-/- versus wild type (WT) YS, but primitive erythroblasts did not circulate in the Ncx1-/- EP. While there was no significant difference in the number of definitive HPCs in Ncx1-/- versus WT YS through E9.5, the Ncx1-/- EP was nearly devoid of HPCs. Thus, primitive erythroblasts and essentially all definitive HPCs destined to initially seed the fetal liver after E9.5 are generated in the YS between E7.0-E9.5 and are redistributed into the EP via the systemic circulation.