Fig. 1

A fluorescent reporter system for real-time measurement of adenosine base-editing activity. a The XMAS-TREE reporter vector consists of a human EF1α promoter driving expression of an mCherry cassette followed by a stop codon (TGA), and a GFP cassette. Targeting pEF-XMAS with an adenine base editor and sg(XMAS) will result in an A-to-G conversion, enabling expression of the downstream GFP reporter. b Two versions of pEF-XMAS-TREE plasmid were designed, one with a single stop codon (XMAS-1xStop) and another with two stop codons (XMAS-2xStop), preceding the coding sequence for GFP. The protospacer sequence (underlined black) for the sgRNA, sg(XMAS), targeting the TGA codon (underlined red) resulting in an A-to-G conversion to TGG and the corresponding amino acid change to tryptophan. The PAM sequence (underlined red) was placed to position the base editing window (underlined orange) around the target nucleotides. c Flow cytometry and d fluorescence microscopy analysis of HEK293 cells at various time points after transfection with pEF-XMAS-1xStop (top panels) or pEF-XMAS-2xStop (bottom panels), pCMV-ABEmax, and sg(XMAS)