Table 3 Frequency of loxP insertion at the Dock7 locus using CRISPR/Cas
Target SiteInjection TypeEmbryos TransferredPups Born/BirthrateNontransgenicLoxp4Loxp5Both Loxp4 and Loxp5Deletion Between Target SitesNull Coat Color
Dock7 cKO1-1Pronuclear1508 (5%)4/8 (50%)2/8 (25%)4/8 (50%)2/8 (25%)4/6 (67%)0/8 (0%)
Dock7 cKO1-2Cytoplasmic15539 (25%)27/39 (69%)8/39 (21%)7/39 (18%)3/39 (8%)15/35 (42%)5/35 (14%)
  • The loxP sites described in the Dock7 cKO1 model, shown in Figure 2, were introduced by either pronuclear or cytoplasmic injection. Mice were screened by genotyping for loxP4, loxP5, deletion of DNA between the Cas9 cut sites (deletion of exon 3–4), and coat color. The number of mice with the indicated genotype or phenotype is listed compared to the total number of mice analyzed. The percentage of mice with the indicated genotype or phenotype is listed in parentheses. Null coat color indicates diluted coat color with the presence of a white belly spot. An example of genotyping results is shown in in Figure 4. Amplification of DNA from tail/toe clips was performed using primer pairs for loxP4 (loxP4A F/R or loxP4B F/R, which provided identical results and could be used interchangeably), loxP5 (loxP5A F/R or loxP5B F/R for loxP5, which provided identical results and could be used interchangeably), and deletion of exons 3–4 (cKO1 ΔB F/R).