CRISPR-Cas systems for genome editing, regulation and targeting

CRISPR-Cas systems for genome editing, regulation and targeting

2014 April ; 32(4): 347–355. doi:10.1038/nbt.2842. | Jeffry D. Sander and J. Keith Joung
The CRISPR-Cas9 system has revolutionized genome editing by enabling precise and efficient modifications of endogenous genes in various cell types and organisms. This technology relies on RNA-guided nucleases (RGNs) to target and cleave specific DNA sequences, leading to either non-homologous end-joining (NHEJ) or homology-directed repair (HDR). The simplicity and high efficiency of this system have facilitated reverse genetics, forward genetic screens, and the creation of large libraries of gRNAs for gene regulation. However, off-target effects remain a significant challenge, and efforts are ongoing to improve specificity through methods such as paired nickases and truncated gRNAs. The CRISPR-Cas9 system also has non-nuclease applications, including gene regulation and labeling of specific genomic loci. Despite these advancements, further improvements are needed to optimize delivery methods, expand targeting range, and enhance HDR efficiency. The potential of CRISPR-Cas9 for therapeutic applications is promising, but challenges related to off-target effects and HDR:NHEJ balance must be addressed.The CRISPR-Cas9 system has revolutionized genome editing by enabling precise and efficient modifications of endogenous genes in various cell types and organisms. This technology relies on RNA-guided nucleases (RGNs) to target and cleave specific DNA sequences, leading to either non-homologous end-joining (NHEJ) or homology-directed repair (HDR). The simplicity and high efficiency of this system have facilitated reverse genetics, forward genetic screens, and the creation of large libraries of gRNAs for gene regulation. However, off-target effects remain a significant challenge, and efforts are ongoing to improve specificity through methods such as paired nickases and truncated gRNAs. The CRISPR-Cas9 system also has non-nuclease applications, including gene regulation and labeling of specific genomic loci. Despite these advancements, further improvements are needed to optimize delivery methods, expand targeting range, and enhance HDR efficiency. The potential of CRISPR-Cas9 for therapeutic applications is promising, but challenges related to off-target effects and HDR:NHEJ balance must be addressed.
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