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Fig. 2
Zbtb11 expression and mne phenotype including delayed neutrophil maturation.
(a) Whole-mount in situ hybridization (WISH) showing widespread expression of Zbtb11 in the developing embryo up until 19 h.p.f., which becomes progressively restricted up until 80 h.p.f. Arrows indicate Zbtb11 expression in the intermediate cell mass (ICM). (b) At 96 h.p.f., mne exhibits ocular, craniofacial and cardiovascular defects. e, eye; h, heart; m, mandibular cartilage. (c) Injection of rhodamine at 48 h.p.f. shows enlarged dye volume in fourth ventricle in mne compared to WT. (d) Loss of rag1 expression in mne at 82 h.p.f. compared to WT. Foxn1 marking the thymic primordium is expressed in mne and WT. (e) RT–qPCR of Zbtb11 expression in FACS sorted adult zebrafish blood cell populations (mean±s.d.; *P≤0.05; n=1 experiment; triplicate replicates on cDNA isolated from purified haemopoietic populations derived from pooled kidney marrows). Ery, erythroid; Lym, lymphoid; Mye, myeloid; Pre, precursors; WKM, whole kidney marrow; Mann–Whitney test. (f) Immunoblot showing ZBTB11 is expressed in human myeloid and lymphoid cell lines and with lower expression in HepG2 hepatocytes (50 μg protein per lane); M, protein ladder with molecular weight in kDa as indicated. (g) Examples of FACS-sorted neutrophils from mne and WT following May–Grünwald Giemsa staining. (h) Quantification of neutrophil sub-populations in mne and WT according to maturity shown as percentage of total cells counted. Schema below graph defines how sub-populations were scored. Gran., granulocytes. n=3 biologically independent experiments (mean±s.e.m.); (a) whole embryo scale70; scale bars, 200 μm (b–d,g).