FIGURE 4.

Generation and genotyping of PWD CD44 allele specific knockout in F1 mice. (A) The distribution of SNPs in Cd44 exons of the whole genome sequenced PWK strain which is a close relative of PWD strain. (B) Two PAM sequences (5′-NGG-3′) that only existed in the PWD mice were selected and confirmed by Sanger sequencing. (C) SgRNA sequences in Cd44 exon5 and exon12 which were selected for allele specific knockout. (D) Workflow of producing PWD CD44 allele specific knockout in F1 mice. (E) Efficiency of CRISPR/Cas9 system in specific inactivation of CD44 from PWD origin. (F) The genotyping of F1-CD44^ASK mice by using capillary electrophoresis analysis (left) and Sanger sequencing (right).