Genome editing using Cas9 nickases
- PMID: 25398340
- DOI: 10.1016/B978-0-12-801185-0.00008-8
Genome editing using Cas9 nickases
Abstract
The RNA-guided, sequence-specific endonuclease Cas9 has been widely adopted as genome engineering tool due to its efficiency and ease of use. Derived from the microbial CRISPR (clustered regularly interspaced short palindromic repeat) type II adaptive immune system, Cas9 has now been successfully engineered for genome editing applications in a variety of animal and plant species. To reduce potential off-target mutagenesis by wild-type Cas9, homology- and structure-guided mutagenesis of Streptococcus pyogenes Cas9 catalytic domains has produced "nicking" enzymes (Cas9n) capable of inducing single-strand nicks rather than double-strand breaks. Since nicks are generally repaired with high fidelity in eukaryotic cells, Cas9n can be leveraged to mediate highly specific genome editing, either via nonhomologous end-joining or homology-directed repair. Here we describe the preparation, testing, and application of Cas9n reagents for precision mammalian genome engineering.
Keywords: CRISPR; Cas9; Gene targeting; Genome editing; Nucleases.
Similar articles
-
Application of CRISPR/Cas9 genome editing to the study and treatment of disease.Arch Toxicol. 2015 Jul;89(7):1023-34. doi: 10.1007/s00204-015-1504-y. Epub 2015 Apr 1. Arch Toxicol. 2015. PMID: 25827103 Review.
-
RNA-dependent DNA endonuclease Cas9 of the CRISPR system: Holy Grail of genome editing?Trends Microbiol. 2013 Nov;21(11):562-7. doi: 10.1016/j.tim.2013.09.001. Epub 2013 Oct 1. Trends Microbiol. 2013. PMID: 24095303
-
Cas9-based genome editing in Arabidopsis and tobacco.Methods Enzymol. 2014;546:459-72. doi: 10.1016/B978-0-12-801185-0.00022-2. Methods Enzymol. 2014. PMID: 25398353
-
Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases and nickases.Genome Res. 2014 Jan;24(1):132-41. doi: 10.1101/gr.162339.113. Epub 2013 Nov 19. Genome Res. 2014. PMID: 24253446 Free PMC article.
-
Expanding the Biologist's Toolkit with CRISPR-Cas9.Mol Cell. 2015 May 21;58(4):568-74. doi: 10.1016/j.molcel.2015.02.032. Mol Cell. 2015. PMID: 26000842 Review.
Cited by 18 articles
-
Upregulated microRNA‑330‑3p promotes calcification in the bicuspid aortic valve via targeting CREBBP.Mol Med Rep. 2020 Sep;22(3):2351-2363. doi: 10.3892/mmr.2020.11297. Epub 2020 Jul 6. Mol Med Rep. 2020. PMID: 32705274 Free PMC article.
-
Applications of Genome Editing Technology in Research on Chromosome Aneuploidy Disorders.Cells. 2020 Jan 17;9(1):239. doi: 10.3390/cells9010239. Cells. 2020. PMID: 31963583 Free PMC article. Review.
-
Advances in Sphingolipidoses: CRISPR-Cas9 Editing as an Option for Modelling and Therapy.Int J Mol Sci. 2019 Nov 24;20(23):5897. doi: 10.3390/ijms20235897. Int J Mol Sci. 2019. PMID: 31771289 Free PMC article. Review.
-
Genome-Wide Off-Target Analysis in CRISPR-Cas9 Modified Mice and Their Offspring.G3 (Bethesda). 2019 Nov 5;9(11):3645-3651. doi: 10.1534/g3.119.400503. G3 (Bethesda). 2019. PMID: 31492696 Free PMC article.
-
CRISPR-Cas9: A multifaceted therapeutic strategy for cancer treatment.Semin Cell Dev Biol. 2019 Dec;96:4-12. doi: 10.1016/j.semcdb.2019.04.018. Epub 2019 May 4. Semin Cell Dev Biol. 2019. PMID: 31054324 Review.
Publication types
MeSH terms
Substances
Grant support
LinkOut - more resources
-
Full Text Sources
-
Other Literature Sources