Author Manuscript Author Manuscript Author Manuscript Author ManuscriptExtended Data Figure E6. Base editing efficiencies of ABE7 variants at 17 genomic sitesNature. Author manuscript; obtainable in PMC 2018 April 25.Gaudelli et al.PageA to G base editing efficiencies in HEK293T cells at 17 human genomic target DNA web pages of ABE7.1-7.five (a), and ABE7.6-7.ten (b). See Extended Information Figure E1 for ABE genotypes and architectures. c, A to G base editing efficiencies in U2OS cells at six human genomic target DNA web-sites of ABE7.8-7.ten. The lower editing efficiencies observed in U2OS cells compared with HEK293T cells are constant with transfection efficiency differences in between the two cell lines; we observed transfection efficiencies of 405 in U2OS cells beneath the situations made use of in this study, when compared with 650 in HEK293T cells. Values and error bars reflect the imply and s.d. of 3 independent biological replicates performed on distinctive days.Author Manuscript Author Manuscript Author Manuscript Author ManuscriptExtended Information Figure E7. Activity window of late-stage ABEsa, Relative A to G base editing efficiencies in HEK293T cells of late-stage ABEs at protospacer positions 1 in two human genomic DNA websites that collectively spot an A at each of those positions.9-cis-Retinoic acid Values are normalized to the maximum observed efficiency at each with the two internet sites for each ABE = 1. b, Relative A to G base editing efficiencies in HEK293T cells of late-stage ABEs at protospacer positions 18 and 20 across all 19 human genomic DNA web pages tested. Values are normalized for the maximum observed efficiency at each of theNature. Author manuscript; offered in PMC 2018 April 25.Gaudelli et al.Page19 web pages for every single ABE = 1. For (a) and (b), values and error bars reflect the mean and s.d. of 3 independent biological replicates performed on diverse days.Author Manuscript Author Manuscript Author Manuscript Author ManuscriptExtended Information Figure E8. Rounds of evolution and engineering increased ABE processivityThe calculated imply normalized linkage disequilibrium (LD) among nearby target As at 6 to 17 human genomic target DNA sites for one of the most active ABEs emerging from each and every round of evolution and engineering. Higher LD values indicate that an ABE is much more likely to edit an A if a nearby A within the identical DNA strand (the identical sequencing study) is also edited. LD values are normalized from 0 to 1 so that you can be independent of editing efficiency. Values and error bars reflect the imply and s.ML115 d. of normalized LD values from 3 independent biological replicates performed on various days.Nature. Author manuscript; obtainable in PMC 2018 April 25.Gaudelli et al.PageAuthor Manuscript Author Manuscript Author ManuscriptExtended Information Figure E9.PMID:23983589 Evaluation of cellular RNAs in ABE7.10-treated cells compared with untreated cellsAuthor ManuscriptRNA from HEK293T cells treated with ABE7.ten as well as the sgRNA targeting website 1, or from untreated HEK293T cells, was isolated and reverse transcribed into cDNA. The cDNAs corresponding to four abundant cellular RNAs (encoding beta-actin, beta-catenin, GAPDH, and RB1), and two cellular RNAs with sequence homology to the tRNA anticodon loop that is the native substrate of E. coli TadA (encoding MN1 and RSLD1), have been amplified and analyzed by HTS. Inside every single amplicon, the mutation frequency at every single adenine position for which the mutation rate was 0.2 is shown for two ABE7.10-treated biologicalNature. Author manuscript; available in PMC 2018 April.
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