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DamID-seq: Genome-wide Mapping of Protein-DNA Interactions by High Throughput Sequencing of Adenine-methylated DNA Fragments
Published on: January 27, 2016
Genome-wide profiling of adenine base editor specificity by EndoV-seq
Puping Liang1,2, Xiaowei Xie1,3, Shengyao Zhi1
1The First Affiliated Hospital, Sun Yat-sen University; MOE Key Laboratory of Gene Function and Regulation, Guangzhou Key Laboratory of Healthy Aging Research, SYSU-BCM Joint Research Center, School of Life Sciences, Sun Yat-sen University, 510275, Guangzhou, China.
Abstract:
The adenine base editor (ABE), capable of catalyzing A•T to G•C conversions, is an important gene editing toolbox. Here, we systematically evaluate genome-wide off-target deamination by ABEs using the EndoV-seq platform we developed. EndoV-seq utilizes Endonuclease V to nick the inosine-containing DNA strand of genomic DNA deaminated by ABE in vitro. The treated DNA is then whole-genome sequenced to identify off-target sites. Of the eight gRNAs we tested with ABE, 2-19 (with an average of 8.0) off-target sites are found, significantly fewer than those found for canonical Cas9 nuclease (7-320, 160.7 on average). In vivo off-target deamination is further validated through target site deep sequencing. Moreover, we demonstrated that six different ABE-gRNA complexes could be examined in a single EndoV-seq assay. Our study presents the first detection method to evaluate genome-wide off-target effects of ABE, and reveals possible similarities and differences between ABE and canonical Cas9 nuclease.
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