alexa Rationally engineered Cas9 nucleases with improved specificity.
Oncology

Oncology

Cancer Medicine & Anti Cancer Drugs

Author(s): Slaymaker IM, Gao L, Zetsche B, Scott DA, Yan WX,

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Abstract The RNA-guided endonuclease Cas9 is a versatile genome-editing tool with a broad range of applications from therapeutics to functional annotation of genes. Cas9 creates double-strand breaks (DSBs) at targeted genomic loci complementary to a short RNA guide. However, Cas9 can cleave off-target sites that are not fully complementary to the guide, which poses a major challenge for genome editing. Here, we use structure-guided protein engineering to improve the specificity of Streptococcus pyogenes Cas9 (SpCas9). Using targeted deep sequencing and unbiased whole-genome off-target analysis to assess Cas9-mediated DNA cleavage in human cells, we demonstrate that "enhanced specificity" SpCas9 (eSpCas9) variants reduce off-target effects and maintain robust on-target cleavage. Thus, eSpCas9 could be broadly useful for genome-editing applications requiring a high level of specificity. Copyright © 2016, American Association for the Advancement of Science.
This article was published in Science and referenced in Cancer Medicine & Anti Cancer Drugs

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