Generation and characterization of developmental and adult hematopoiesis of gata2b mutant zebrafish. (A) Schematic of the gata2b gene with 6 exons. CRISPR gRNA (red triangle) targets in exon 2. KO, gata2b^ko; NRD, negative regulatory domain; TAD, transactivating domain; WT, wild-type; ZF, zinc finger. (B) CRISPR edit in gata2b^ko zebrafish at position C10. Dotted line = gRNA sequence; solid line = PAM site. Predicted gata2b mutant protein shown. (C) In situ hybridizations of embryos from gata2b^het in cross for runx1 at 24 hpf (n = 3 clutches), and cmyb at 72 hpf (n = 4 clutches). Bar graphs quantify proportion of embryos (mean ± standard deviation) in blindly scored categories shown in representative images; areas of interest marked by black arrowheads. Number of embryos in each genotype is shown. *P < .02 by unpaired 2-tailed Student t test, gata2b^het (high) vs gata2b^ko (high); n.s., not significant. (D) Quantification of marrow myelomonocytes in WT, gata2b^het, and gata2b^ko zebrafish. *P < .03 compared with WT and/or gata2b^het, 1-way ANOVA. WT, n = 18, 19, and 23; gata2b^het, n = 76, 41, and 75; gata2b^ko, n = 48, 23, and 51, at 3 months, 8 months, and 12 months, respectively. (E) Relative ratio of progenitors to myelomonocytes in the marrow at 8 months. *P < .02, unpaired 2-tailed Student t test, n = 19 WT, n = 41 gata2b^het, n = 23 gata2b^ko. (F) Quantification of myelomonocytes in the donor cells of whole marrow cell transplantation recipients. Recipients with at least 10% donor engraftment were analyzed. *P < .0001 for WT or gata2b^het vs gata2b^ko, 1-way ANOVA, n = 20 WT, n = 29 gata2b^het, n = 37 gata2b^ko recipients.