Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases
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TLDR
A novel algorithm termed Cas-OFFinder that searches for potential off-target sites in a given genome or user-defined sequences and allows variations in protospacer-adjacent motif sequences recognized by Cas9, the essential protein component in RGENs.Abstract:
Summary: The Type II clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system is an adaptive immune response in prokaryotes, protecting host cells against invading phages or plasmids by cleaving these foreign DNA species in a targeted manner. CRISPR/Cas-derived RNA-guided engineered nucleases (RGENs) enable genome editing in cultured cells, animals and plants, but are limited by off-target mutations. Here, we present a novel algorithm termed Cas-OFFinder that searches for potential off-target sites in a given genome or user-defined sequences. Unlike other algorithms currently available for identification of RGEN off-target sites, Cas-OFFinder is not limited by the number of mismatches and allows variations in protospacer-adjacent motif sequences recognized by Cas9, the essential protein component in RGENs. Cas-OFFinder is available as a command-line program or accessible via our website.
Availability and implementation: Cas-OFFinder free access at http://www.rgenome.net/cas-offinder.
Contact: rk.ca.uns@uaseab or rk.ca.uns@10miksjread more
Citations
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Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9
John G. Doench,Nicolo Fusi,Meagan E Sullender,Mudra Hegde,Emma W Vaimberg,Katherine F Donovan,Ian Smith,Zuzana Tothova,Zuzana Tothova,Craig B. Wilen,Robert C. Orchard,Herbert W. Virgin,Jennifer Listgarten,David E. Root +13 more
TL;DR: Recently devised sgRNA design rules are used to create human and mouse genome-wide libraries, perform positive and negative selection screens and observe that the use of these rules produced improved results, and a metric to predict off-target sites is developed.
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TL;DR: In vivo generation of mouse models carrying clinically relevant mutations using C→T and A→G editors is demonstrated, making it feasible to model and potentially cure relevant genetic diseases.
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High-fidelity CRISPR–Cas9 nucleases with no detectable genome-wide off-target effects
Benjamin P. Kleinstiver,Vikram Pattanayak,Michelle S. Prew,Shengdar Q. Tsai,Nhu T. Nguyen,Zongli Zheng,J. Keith Joung +6 more
TL;DR: With its exceptional precision, SpCas9-HF1 provides an alternative to wild-type Sp Cas9 for research and therapeutic applications and suggests a general strategy for optimizing genome-wide specificities of other CRISPR-RNA-guided nucleases.
In vivo genome editing using Staphylococcus aureus Cas9
F. Ann Ran,Le Cong,Winston X. Yan,David A. Scott,Jonathan S. Gootenberg,Andrea J. Kriz,Bernd Zetsche,Ophir Shalem,Xuebing Wu,Kira S. Makarova,Eugene V. Koonin,Phillip A. Sharp,Feng Zhang +12 more
TL;DR: In this paper, the RNA-guided endonuclease Cas9 has emerged as a versatile genome-editing platform and has been used for basic research and therapeutic applications that use the highly versatile adeno-associated virus (AAV) delivery vehicle.
Journal ArticleDOI
In vivo genome editing using Staphylococcus aureus Cas9
F. Ann Ran,Le Cong,Winston X. Yan,David A. Scott,Jonathan S. Gootenberg,Andrea J. Kriz,Bernd Zetsche,Ophir Shalem,Xuebing Wu,Kira S. Makarova,Eugene V. Koonin,Phillip A. Sharp,Feng Zhang +12 more
TL;DR: Six smaller Cas9 orthologues are characterized and it is shown that Cas9 from Staphylococcus aureus (SaCas9) can edit the genome with efficiencies similar to those of SpCas9, while being more than 1 kilobase shorter.
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