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Fig. 4 | BMC Biotechnology

Fig. 4

From: Enhancement of CRISPR-Cas9 induced precise gene editing by targeting histone H2A-K15 ubiquitination

Fig. 4

Fluorescence-based DSB repair assay using Rad18UBD and RNF169UBD fusion proteins in HEKTLR6 reporter cells. Fusion constructs for BRCA1 or TetR with Rad18UBD or RNF169 UBD were cotransfected with the TLR HDR repair template (TLR-donor-tetO), sgRNA and Cas9 into HEKTLR6 cells. The frequency of Venus+ cells (green bars) and RFP+ cells (red bars) within the population of BFP+ cells was measured by FACS analysis 72 h after transfection and used to calculate the ratio of Venus/RFP positive cells. The X-axis shows the transfected samples and the selection of cotransfected plasmids below. Samples 1 and 2 are controls showing the basic frequency of Venus+ and RFP+ cells upon transfection with Cas9 and sgRNA or in combination with TLR-donor-tetO as repair template. Data from four independent experiments, each with three replicates per sample, are represented as mean values ± SD. Statistical significance of samples 3–16 in comparison to the control sample 2 was determined by two-way ANOVA and Dunnett’s multiple comparison tests with **P < 0.01, ***P < 0.001 (HDR) and ##P < 0.01, ###P < 0.001 (NHEJ). Raw data are shown in the Supplementary data file

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