Generation of Kif15 mutant alleles using CRISPR/Cas9 in AB line zebrafish.
(A) Target site k7 and k8 were shown to be effectively cleaved in AB line zebrafish, forming chimera, and various heritable mutations were found in the F1 generation. Underlined sequences are proto-spacers, while Double underlined triplets are PAM.
(B) Sequence alignment of wildtype and inherited mutated alleles of kif15. Underlined fragment is the proto-spacer identified by sgRNA k7. Double underlined triplets are PAM recognized by Cas9.
(C) Deduced translation products showed that mutated kif15 alleles code for disrupted Kif15 proteins lacking part of the kinesin motor domain and the entire tail. Numbers indicate peptide length. The numbers 216 and 219 indicate loss of residues 217 and 218 in mutated allele ntu203.