The establishment of the hematopoietic stem cell (HSC) pool in vertebrates depends not only on the formation and the propagation of these stem cells but also on their proper trafficking among the defined hematopoietic organs. However, the physiological mechanisms that regulate HSC mobilization remain elusive. Through analysis of the zebrafish cmyb mutant cmybhkz3, we show that the suppression of cMyb function abrogates larval and adult hematopoiesis, with concomitant accumulation of hematopoietic stem/progenitor cells (HSPCs) in their birthplace, the ventral wall of the dorsal aorta (VDA). Cell tracking and time-lapse recording reveal that the accumulation of HSPCs in cmybhkz3 mutants is caused by the impairment of HSPC egression from the VDA. Further analysis demonstrates that the HSPC migratory defects in cmybhkz3 mutants are at least partly due to adversely elevated levels of chemokine stromal cell-derived factor 1a (Sdf1a). Our study reveals that cMyb plays a hitherto unidentified role in dictating physiological HSPC migration by modulating Sdf1a signaling.