Fig. 3: Programmable DNA ADP-ribosylation primarily drives base mutagenesis in yeast and plants.
a, Experimental setup for introducing a six-base replacement with two adjacent premature stop codons in the FCY1 gene of S. cerevisiae. NLS, nuclear localization signal. b, Impact of ADPr-TAE on templated recombination in the presence of an RT. Bars and error bars represent the mean and s.d. of three independent experiments started from separate transformations. Dots represent individual measurements. c, Impact of ADPr-TAE on mutagenesis of the ADP-ribosylated thymine in the presence or absence of an RT. d, Frequency of base mutations across the sgRNA target. Each black bar specifies DarT2 recognition motifs, while the red base specifies the ADP-ribosylated base within the motif. Representative Sanger sequencing chromatograms can be found in Extended Data Fig. 6a,b. In c and d, bars and error bars represent the mean and s.d. of three independent experiments started from separate transformations. e, Experimental setup for assessing ADPr-TAE without an RT in N. benthamiana. f, Frequency of base mutagenesis of the ADP-ribosylated thymine in the sgRNA1 target in the PDS1 gene compared to the NT control. g, Frequency of base mutations across the DNA target for sgRNA1–sgRNA3 compared to the NT control. The location of base mutations under targeting and non-targeting conditions can be found in Supplementary Fig. 6. In f and g, bars and error bars represent the mean and s.e.m. of three independent experiments started from separate transformations.
