Fig. 1

Generation of Irx1 knock-out zebrafish. Targeted gene mutation was performed by TALEN and CRISPR/Cas9 knockout methods for Irx1a (A) and for Irx1b gene (B), respectively. Frameshift mutation was caused by the knockout strategies. Insertion or deletion of the nucleotides was indicated by red letterings. (C) Three-day-old embryos of the zebrafish mutants. While either Irx1aor Irx1b homozygote mutants did not show abnormal morphology during embryonic development, some siblings (homozygote for both Irx1a and Irx1b) from the breeding of Irx1a^−/−/1b^+/− and Irx1a^+/−/1b^−/− showed severe morphologic abnormality. This suggests that majority of homozygote mutants of Irx1a and Irx1b are developmentally defective. A few of Irx1a^−/−/1b^−/− zebrafish, however, were found to survive up to 3 months of age but were not fertile and died within 6 months of age.