Computational analysis GeCKO v2 library (human)

Table 1 - Non-specific sgRNAs GeCKO v2 (human). This table represents the total of 5664 sgRNAs within the entire human GeCKO v2 library (half-libraries A and B combined) that recognise at least two identical target sites (20 bp protospacer + NGG PAM) within the reference genome. For the analysis, sgRNA target sites were aligned against the human reference genome (GRCh38/hg19 – primary assembly) using BLAT (v36) after optimisation of the algorithm for short length queries (-tileSize=6, -stepSize=3). The resulted alignments were further filtered to only include perfect matches and unique targets (exclusion of hits on alternative assemblies). Analysis was performed using R (v3.2.3) in R studio (v1.0.136) using Ubuntu.

Table 2 - Flagged genes GeCKO v2 (human). This table represents the total of 775 genes for which more than half the corresponding sgRNAs within the entire human GeCKO v2 library (half-libraries A and B combined) are non-specific, i.e. recognise at least two identical target sites (20 bp protospacer + NGG PAM) within the reference genome. A threshold of >50% non-specific sgRNAs per gene (e.g. 4 out of 6 for protein coding genes; 3 out of 4 for miRNA genes) was set in order to flag a group of sgRNAs. Analysis was performed using R (v3.2.3) in R studio (v1.0.136) using Ubuntu.