Fig. 1: PSCs-derived epidermis and appendages generated on chimeric dermis by p63 knockout of host embryos.

a CRISPR-Cas9 gRNA design for p63 gene. b Macroscopic images of a WT (left) mouse and a p63 knockout mouse (right). Scale bars, 5 mm. c Experimental schematic for p63 knockout chimera generation (E9.5 to E18.5 by C-section). Created with BioRender.com. d Flow cytometry of p63 knockout chimera epidermis (E18.5). e The comparison of chimerism between WT chimera and p63 knockout chimera (Splenocyte; CD45⁺, Melanocyte; CD45⁻CD117⁺, Keratinocyte; CD45⁻CD117⁻CD49f⁺). Statistical analysis was performed using two-sided Friedman tests followed by Dunn’s test with correction for multiple comparisons. Square, WT chimera (n = 13); triangle, p63 knockout chimera (n = 15). f Macroscopic and fluorescence (EGFP) images of four p63 knockout chimera (E18.5) with different global chimerism (splenocytes). The upper number is skin-covered areas, and the lower number is global chimerism in each chimera. Scale bars, 5 mm. g Histological staining of WT, p63 knockout, and p63 knockout chimera skin. Scale bar, 100 μm. h Immunofluorescence staining of the skin of p63 knockout chimera. DAPI, nuclear; EGFP, the label of donor cell; p63, basal cell marker; KRT14, mature keratinocytes in the basal layer. Scale bars, 50 μm. i Magnified hair follicle image of yellow box in (h). Scale bars, 20 μm. Source data are provided with this paper.