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Fig. 3 | BMC Biology

Fig. 3

From: BE-FLARE: a fluorescent reporter of base editing activity reveals editing characteristics of APOBEC3A and APOBEC3B

Fig. 3

Co-enrichment of editing events using BE-FLARE is superior to selecting transfected cells with BE3-T2A-RFP. a Schematic diagram of the enrichment strategy for genomic co-editing events using the BE-FLARE reporter. b Co-selection with transient expression of BE-FLARE. PC9 cells were co-transfected with BE-FLARE and a plasmid expressing BE3 and gRNAs targeting BFP and either BRAF T57/Q58 (left) or EGFR T790 (right). Cells were mock-sorted (all viable cells) or sorted for GFP expression by FACS 72 h after transfection and grown for 120 h before harvesting genomic DNA for Sanger sequencing. Cells transfected with non-targeting gRNA served as a negative control reference. Percentage editing was estimated from sequencing chromatograms using EditR [31]. Data are expressed as the mean ± SD of two independent experiments. c Schematic of the methodology used to monitor base editing efficiency and BE3 expression using a BE3-T2A-TurboRFP construct (BE3-RFP). d BE-FLARE is superior to BE3-T2A-TurboRFP in marking BE3-active cells. PC9 cells stably expressing BE-FLARE were transfected with BE3 or BE3-T2A-turboRFP. Cells were analysed by flow cytometry 72 h after transfection to quantify GFP (edited) and RFP-expressing cells. Data are representative of two independent experiments. Raw data can be found in Additional file 2

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