Fig. 4.

Fig. 4.

Allele exchange in the obelix gene. In A the start of the coding sequence for obe is shown. The CRISPR target site is indicated by an arrow, the two point mutations introduced by HDR are shown in red and green, respectively, the recognition site for SalI that is generated by the HDR is underlined. The oligonucleotide used as donor DNA is also depicted. In B sequences from wild-type fish, and fish heterozygous or homozygous for the engineered allele are shown, the two positions that were engineered are boxed. There are no discernible differences between wild-type siblings (C) and fish heterozygous for the engineered allele (kcnj13^ten/+) (E), nor between fish that carry a kcnj13 loss-of-function allele over a wild-type allele (kcnj13^t24ui/+) (F) or over the engineered allele (kcnj13^t24ui/ kcnj13^ten) (G). For comparison, a fish homozygous mutant for a loss-of-function allele of obe (kcnj13^t24ui/kcnj13^t24ui) is shown in D, scale bar: 1 cm.