Supplementary Figure 1: Characterization of AID variants.
From: Directed evolution using dCas9-targeted somatic hypermutation in mammalian cells
a) Diagram of AID variants. NLS, NES, deaminase domain, truncations, and activity-altering mutations are indicated. b) Fluorescence microscopy of MS2-AID and MS2-AIDΔ constructs in K562 cells is shown. Cells were fixed and stained with an MS2 antibody (green) and the nuclear stain DAPI (blue). Images shown are representative of four collected images. c) A comparison of the expression of different MS2-AID variants is shown. K562 cells expressing the variants were lysed and analyzed on an SDS-PAGE gel followed by immunoblotting with an MS2 antibody (top) or GAPDH antibody (bottom). d) K562 cells containing dCas9, GFP, and mCherry were transiently electroporated with indicated combinations of MS2-AID, MS2-AIDΔ, or MS2-AIDΔDead and either sgGFP.1 or sgNegCtrl. GFP and mCherry fluorescence of the cells were measured by flow cytometry as a proxy for mutation rate. Shown are the scatter plots from the flow cytometry and a graph summarizing the non-fluorescent populations. e) Cells were sorted for low GFP expression and the GFP locus was sequenced. A graph of the enrichment of mutation at each base is shown here.
