CDC STACKS serves as an archival repository of CDC-published products including scientific findings, journal articles, guidelines, recommendations, or other public health information authored or co-authored by CDC or funded partners.
As a repository, CDC STACKS retains documents in their original published format to ensure public access to scientific information.
i
Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity
-
Aug 29 2013
-
-
Source: Cell. 2013; 154(6):1380-1389.
Details:
-
Alternative Title:Cell
-
Personal Author:
-
Description:Targeted genome editing technologies have enabled a broad range of research and medical applications. The Cas9 nuclease from the microbial CRISPR-Cas system is targeted to specific genomic loci by a 20 nt guide sequence, which can tolerate certain mismatches to the DNA target and thereby promote undesired off-target mutagenesis. Here, we describe an approach that combines a Cas9 nickase mutant with paired guide RNAs to introduce targeted double-strand breaks. Because individual nicks in the genome are repaired with high fidelity, simultaneous nicking via appropriately offset guide RNAs is required for double-stranded breaks and extends the number of specifically recognized bases for target cleavage. We demonstrate that using paired nicking can reduce off-target activity by 50- to 1,500-fold in cell lines and to facilitate gene knockout in mouse zygotes without sacrificing on-target cleavage efficiency. This versatile strategy enables a wide variety of genome editing applications that require high specificity.
-
Subjects:
-
Source:
-
Pubmed ID:23992846
-
Pubmed Central ID:PMC3856256
-
Document Type:
-
Funding:
-
Volume:154
-
Issue:6
-
Collection(s):
-
Main Document Checksum:
-
Download URL:
-
File Type:
[PDF-1.36 MB]
