ZFN-mediated targeted mutagenesis of cbfb. (A) A schematic of genomic organization of cbfb with numbered boxes depicting exons, with connecting lines depicting introns and red bars marking the location of the ZFN pair used to generate knockout mutants. (B) Alignment of nucleotide sequences from nt198-231 of the cbfb open reading frame to show the ZFN target site (gray highlight in WT), spacer sequence (red letters), and exact sequences of the mutant alleles, del4 and ins4, which are characterized in this study. Yellow highlighted area marks the deleted or inserted nucleotides. (C) At 36 hpf, expression of cbfb in WT sibling was detectable in the ventral DA, where HSCs originate (black arrowhead). cbfb expression in the ventral DA was abrogated in both cbfb^del4/del4 (D) and cbfb^ins4/ins4 mutants (E).