Identification of high-efficiency 3’GG gRNA motifs in indexed FASTA files with ngg2
- Published
- Accepted
- Subject Areas
- Bioinformatics, Computational Biology
- Keywords
- gRNA, motif discovery, python, open-source, CRISPR/Cas9, 3'GG
- Copyright
- © 2015 Roberson
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ PrePrints) and either DOI or URL of the article must be cited.
- Cite this article
- 2015. Identification of high-efficiency 3’GG gRNA motifs in indexed FASTA files with ngg2. PeerJ PrePrints 3:e969v2 https://doi.org/10.7287/peerj.preprints.969v2
Abstract
CRISPR/Cas9 is emerging as one of the most-used methods of genome modification in organisms ranging from bacteria to human cells. However, the efficiency of editing varies tremendously site-to-site. A recent report identified a novel motif, called the 3’GG motif, which substantially increases the efficiency of editing at all sites tested in C. elegans. Furthermore, they highlighted that previously published gRNAs with high editing efficiency also had this motif. I designed a python command-line tool, ngg2, to identify 3’GG gRNA sites from indexed FASTA files. As a proof-of-concept, I screened for these motifs in six model genomes: Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, Mus musculus, and Homo sapiens. I also scanned the genomes of pig (Sus scrofa) and African elephant (Loxodonta africana) to demonstrate the utility in non-model organisms. I identified more than 60 million single match 3’GG motifs in these genomes. Greater than 61% of all protein coding genes in the reference genomes had at least one unique 3’GG gRNA site overlapping an exon. In particular, more than 96% of mouse and 93% of human protein coding genes have at least one unique, overlapping 3’GG gRNA. These identified sites can be used as a starting point in gRNA selection, and the ngg2 tool provides an important ability to identify 3'GG editing sites in any species with an available genome sequence.
Author Comment
This updated version of the manuscript reflects changes made to the paper after comments returned from review. The tool has since been updated to allow multi-processing and removes external dependencies on samtools. Two additional non-model species are included to demonstrated utility, and the scan has been updated to exhaustive genome search rather than a block-like scan that would only find one gRNA per 20bp window, ignoring any additional overlapping gRNAs.