cMyb regulates HSPC migration through modulating Sdf1a level. (A) Double staining of Cd41:eGFP protein (top panel) and sdf1a RNA (middle panels) in 53-hpf siblings (left panel) and cmyb^hkz3 mutant embryos (right panel) of cmyb^hkz3/Tg(cd41:eGFP) (representative figures). The bottom panels show merged images. (B) Quantitative RT-PCR for sdf1a gene expression in purified Cd41-eGFP⁺ cells from the VDA region of cmyb^hkz3/Tg(cd41:eGFP) mutants and siblings. Units on the y-axis represent the relative fold change of sdf1a expression in siblings and cmyb^hkz3 mutant embryos. Expression level was normalized with β-actin expression. Error bars, SD. (C) HSPC migratory defect in cmyb^hkz3 was rescued by sdf1a MO knockdown. cmyb^hkz3/Tg(cd41:eGFP) mutant and sibling embryos were injected with sdf1a MO plus DMNB-caged Flu (blue columns) or control MO plus DMNB-caged Flu (yellow columns) at 1-cell stage and cells in the aortic floor were then labeled through photoactivation. Emergence of eGFP/Flu double-positive cells in the CHT (gray numbers in each column) were scored and quantified 16 hours after photoactivation. Units on the y-axis represent the average number of eGFP/Flu double-positive cell per embryo (n ≥ 19, mean ± SE). An asterisk indicates a statistics significant increase of the number of eGFP/Flu double-positive cells migrated to the CHT in cmyb^hkz3 mutants injected with sdf1a MO compared with cmyb^hkz3 mutants injected with control MO (t test, P < .05).