SpCas9 activity prediction by DeepSpCas9, a deep learning–based model with high generalization performance

To increase the accuracy of the analysis, deep sequencing data were filtered; target sequences with deep sequencing read counts below 200 and background indel frequencies above 8% were excluded as similarly performed previously (21). DNase-sequencing (DNase-seq) narrow peak data from ENCODE (36) were used to calculate chromatin accessibility as previously described (21). For each target site, 23 bases of the PAM plus protospacer sequence were aligned to the hg19 human reference genome using bowtie (41). Only the target sites that overlapped with DNase-seq narrow peaks were considered as DNase I hypersensitive target sites. We divided the Endo_Cas9 dataset into paired subsets by stratified random sampling from strata of DHS and non-DHS sites so that a similar ratio of DHS/non-DHS sites was assigned to each subset.