Zebrafish rb1Δ7/Δ7 homozygous mutant larval brain transcriptome and neurogenic phenotype. (A) Zebrafish rb1/rb1 mutant transcriptome was generated from rb1Δ7/Δ7 homozygous and +/+ wild-type siblings from a cross between heterozygous rb1Δ7/+ adults. Heads from 5 dpf larva were dissected and trunk tissue genotyped. Three pools of five heads of each genotype were used to generate RNA-Seq libraries. (B,C) Wild-type (B) and rb1Δ7/Δ7 (C) 5 dpf larvae. (D) Diagram of larval midbrain and retina with location of neural stem and progenitor cells at brain ventricle (V) in the optic tectum (OT) and thalamic region (Th), and at the retina ciliary marginal zone (cmz). Mature, postmitotic neurons are located in the lateral brain parenchyma and inner retina. (E-L) Immunolocalization with neural differentiation marker HuC/D and mitotic M-phase marker phosphohsitone H3 (E,F,I,J). Wild-type brain and retina show the organization of mature neurons in dorsal brain optic tectum (OT), ventral brain thalamus (Th), and laminated layers of the retina (gcl, ganglion cell layer; inl, inner nuclear layer; onl, outer nuclear layer). Only one mitotic cell is detected at the brain ventricle (F, arrow). Mutant brain shows M-phase cells scattered throughout the midbrain tectum and thalamus (G,H). In the retina, numerous M-phase cells are present across the entire inner nuclear layer, with occasional cells in the outer nuclear and ganglion cell layer (K,L). (M,N) Quantification of pH3-positive cells in midbrain (M) and retina (N) sections from +/+ and rb1Δ7/Δ7 individual 5 dpf larvae (n=16 for each genotype). Data are mean±s.e.m. P-values calculated by two-tailed unpaired Student's t-test. Scale bars: 200 µm (B,C); 50 µm (E,I,G,K); 20 µm (F,J,H,L).
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