Fig. 1

Generation of epoa mutant zebrafish by genomic editing using CRISPR/Cas9. (A) A 11-bp deletion and 0-bp addition in epoa exon 2 were identified in CRISPR/Cas9 generated mutants, which predicted a stop codon. (B) Schematic representation of the three epoa transcripts, with the position of the Cas9 target in red and the new stop codon appearing after deletion of 11 bp in black. (C) DNA agarose gel electrophoresis to distinguish WT, epoaszy8 and epoa+/szy8. (D) Protein structure prediction shows that the main erythropoietin (EPO)-THPO domain is completely truncated. (E and F) epoa?11,+0 mutated transcripts generated in epoaszy8 mutants. Primer pairs WT-FP/CO-RP, Mut-FP/CO-RP and CO-FP/CO-RP were used to detect WT and epoaszy8 genotypes, respectively. Statistical significance was determined using a two sample Student's t-test, n ? 15 per group, and data were combined from four replicates, mean ± SEM, nd: no detective, *p < .05, **p < .001 and ***p < .001. (G) Survival curves of the epoa mutant and its siblings. (H) Percentage of epoa+/+, epoa+/? and epoa?/? zebrafish survival at 5 dpf, 1 mpf and 2.5 mpf (n = 27?120 embryos/larvae/fish).