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CRISPR-Cas systems for editing, regulating and targeting genomes
Jeffry D Sander1,2 & J Keith Joung1,2
Targeted genome editing using engineered nucleases has rapidly gone from being a niche technology to a mainstream method used by many biological researchers. This widespread adoption has been largely fueled by the emergence of the clustered, regularly interspaced, short palindromic repeat (CRISPR) technology, an important new approach for generating RNA-guided nucleases, such as Cas9, with customizable specificities. Genome editing mediated by these nucleases has been used to rapidly, easily and efficiently modify endogenous genes in a wide variety of biomedically important cell types and in organisms that have traditionally been challenging to manipulate genetically. Furthermore, a modified version of the CRISPR-Cas9 system has been developed to recruit heterologous domains that can regulate endogenous gene expression or label specific genomic loci in living cells. Although the genome-wide specificities of CRISPR-Cas9 systems remain to be fully defined, the power of these systemsto perform targeted, highly efficient alterations of genome sequence and gene expression will undoubtedly transform biological research and spur the development of novel molecular therapeutics for human disease.
npg 201 4 Nature America, Inc. All rights reserved.
The introduction of targeted genomic sequence changes into living cells and organisms has become a powerful tool for biological research and is a potential avenue for therapy of genetic diseases. Frameshift knockout mutations enable reverse genetics and identification of gene functions; sequence insertions can fuse epitope tags or other functional domains, such as fluorescent proteins, to endogenous gene products; and specific sequence alterations can induce amino acid substitutions for disease modeling, transfer traits in agricultural crops and livestock, and correct defective genes for therapeutic applications. For many years, strategies for efficiently inducing precise, targeted genome alterations were limited to certain organisms (e.g., homologous recombination in yeast or recombineering in mice) and often required drug-selectable markers or left behind scar sequences associated with the modification method (e.g., residual loxP sites from Cre recombinase-mediated excision). Targeted genome editing using customized nucleases provides a general method for inducing targeted deletions, insertions and precise sequence changes in a broad range of organisms and cell types. The high efficiency of genome editing obviates the need for additional sequences, such as drug-resistance marker genes,...