Lee, Ciaran M.Cradick, Thomas J.Bao, Gang2017-05-042017-05-042016Lee, Ciaran M., Cradick, Thomas J. and Bao, Gang. "The Neisseria meningitidis CRISPR-Cas9 System Enables Specific Genome Editing in Mammalian Cells." <i>Molecular Therapy,</i> 24, no. 3 (2016) Elsevier: 645-654. https://doi.org/10.1038/mt.2016.8.https://hdl.handle.net/1911/94171The clustered regularly-interspaced short palindromic repeats (CRISPR)—CRISPR-associated (Cas) system from Streptococcus pyogenes (Spy) has been successfully adapted for RNA-guided genome editing in a wide range of organisms. However, numerous reports have indicated that Spy CRISPR-Cas9 systems may have significant off-target cleavage of genomic DNA sequences differing from the intended on-target site. Here, we report the performance of the Neisseria meningitidis (Nme) CRISPR-Cas9 system that requires a longer protospacer-adjacent motif for site-specific cleavage, and present a comparison between the Spy and Nme CRISPR-Cas9 systems targeting the same protospacer sequence. The results with the native crRNA and tracrRNA as well as a chimeric single guide RNA for the Nme CRISPR-Cas9 system were also compared. Our results suggest that, compared with the Spy system, the Nme CRISPR-Cas9 system has similar or lower on-target cleavage activity but a reduced overall off-target effect on a genomic level when sites containing three or fewer mismatches are considered. Thus, the Nme CRISPR-Cas9 system may represent a safer alternative for precision genome engineering applications.engThis work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material.Journal articlehttps://doi.org/10.1038/mt.2016.8