Fig. 4: Targeting strategies for specifically disrupting dominant alleles of Fgfr3^G369C/+ mice.

a Schematic of the dominant-negative effects exerted by dominant mutations and correction strategy by Past-CRISPR. Dominant mutations (red dot) do lead to the nonmutational impairment of the wild-type allele (black dashed line). Dominant mutation allele could be separated as the isolation of parental genome at the zygote pronuclear stage and disrupted by Past-CRISPR. b Percentage of induced mutation types in exon 4 of Fgfr3 by Past-CRISPR for each of two sgRNAs. The target genotype of each embryo was analyzed by TIDE. Two independent experiments were performed for every sgRNA. Source data are provided as a Source data profile. c TA cloning sequences resulting from the tail of one of the Fgfr3^{G369C del/+} mice. The dominant mutation site is marked by the right red box. The indels induced by Fgfr3 sgRNA1 are marked by the left red box. A total of three Fgfr3^{G369C del/+} mice were tested and 20 TA clones were sequenced in each sample. d Morphology of 2-month-old Fgfr3^G369C/+ mice (left), wild-type mice (middle), and Fgfr3^{G369C del/+} mice (right). e Microcomputed tomography analyses show the skeletons of wild-type (WT), Fgfr3^{G369C del/+}, and Fgfr3^G369C/+ mice. f Microcomputed tomography analyses on the maxillofacial bone. Skulls and jaws are significantly reduced in size and mainly exhibit severe dysplasia in Fgfr3^G369C/+ mice compared with wild-type mice. The bone contours have been corrected to normal in Fgfr3^{G369C del/+} mice. g Trabecular bone quantification analyses of femurs from wild-type (WT), Fgfr3^{G369C del/+}, and Fgfr3^G369C/+ mice. Femurs of Fgfr3^G369C/+ mice exhibit cortical bone thickness, noticeably sparse bone trabecular and shortened backbone. Femurs of Fgfr3^{G369C del/+} mice are corrected to nearly normal levels. h Quantitative micro-CT analysis of distal femoral metaphysis from wild-type (WT), Fgfr3^{G369C del/+}, and Fgfr3^G369C/+ mice. Results are expressed as the mean ± SEM. of six technical replicates per group. P value (one-way ANOVA) are also shown. Source data are provided as a Source data profile. i Undecalcified bone section analysis of femurs shows primary sparse bone trabecula and secondary ossification centers of Fgfr3^G369C/+ mice and normal states in Fgfr3^{G369C del/+} mice when compared to that of wild-type mice.