Fig. 3

TET3 knockdown impairs HR-mediated DNA repair efficacy in vitro in mCFs. A–B mCFs integrated with a DR-GFP HR reporter substrate were transfected with TET3 knockdown construct and I-SceI and analyzed for change in HR efficiency by scoring % of GFP/RFP double-positive cells using flow cytometry. The associated graph represents HR efficacy in the ratio % of GFP/RFP double-positive cells. C–D mCFs integrated with a pLCN-DSB NHEJ reporter substrate were transfected with TET3 knockdown construct and I-SceI and analyzed for change in NHEJ efficiency by scoring % of GFP/RFP double-positive cells using flow cytometry. E Bar graph representing a decrease in BrdU incorporation under non-denaturing condition, a direct readout for DNA end resection in TET3 knockdown mCFs which can be rescued upon TET3 overexpression. F Western blot confirming TET3 knockdown and rescue in mCFs. G and H Representative confocal images and an associated graph shows rad51 (green) and γH2AX (red) co-localization (yellow) in mCFs in control, TET3 knockdown, and rescued cells (n = 100 cells were analyzed in each condition from 3 different experiments). For comparing between two groups two-tailed Student’s t test was performed. For comparison between more than two groups, statistical significance was calculated using one-way ANOVA parametric analysis, n.s. represents non-significant and P values correspond to *p ≤ 0.05, **p ≤ 0.01, ***P ≤ 0.01