Fig. 3

Cellular characteristics of canine fibroblasts immortalized by CRISPR/Cas9-mediated TP53 knockout (KO). a No enhanced green fluorescent protein (EGFP) expression was detected by fluorescence activated cell sorting (FACS) analysis in both TP53KO#30 and TP53KO#39 cell lines. b Relative proliferation rates of each cell lines (× 15,000 cells). c Cumulative growth curve data of the control (early passage 2) and two TP53 KO cells were obtained after additional 14 consecutive passages in culture. d Representative images showing the cellular morphologies of the control and two TP53 KO cells at the end time-point (passage 14) of cumulative growth counting analysis. e Representative images of the control and two TP53 KO cells stained with senescence associated β-galactosidase (SA-β-gal) at the end time-point of cumulative growth counting analysis. f Percentage of SA-β-gal-positive cells in each group. g Immunoblot data showing TP53, p21, and SV40LT protein levels in the control, SV40LT-transduced, TP53KO#30, and TP53KO#39 cells grown in the absence or presence of 100 μM TMZ for 48 h. α-Tubulin was used as the loading control. **, p < 0.01