Protein Engineering Strategies to Expand CRISPR-Cas9 Applications

International Journal of Genomics
Lucas F RibeiroRichard J Ward

Abstract

The development of precise and modulated methods for customized manipulation of DNA is an important objective for the study and engineering of biological processes and is essential for the optimization of gene therapy, metabolic flux, and synthetic gene networks. The clustered regularly interspaced short palindromic repeat- (CRISPR-) associated protein 9 is an RNA-guided site-specific DNA-binding complex that can be reprogrammed to specifically interact with a desired DNA sequence target. CRISPR-Cas9 has been used in a wide variety of applications ranging from basic science to the clinic, such as gene therapy, gene regulation, modifying epigenomes, and imaging chromosomes. Although Cas9 has been successfully used as a precise tool in all these applications, some limitations have also been reported, for instance (i) a strict dependence on a protospacer-adjacent motif (PAM) sequence, (ii) aberrant off-target activity, (iii) the large size of Cas9 is problematic for CRISPR delivery, and (iv) lack of modulation of protein binding and endonuclease activity, which is crucial for precise spatiotemporal control of gene expression or genome editing. These obstacles hinder the use of CRISPR for disease treatment and in wider biotechnolog...Continue Reading

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Citations

Feb 6, 2019·BioMed Research International·Lucas F RibeiroMaría-Eugenia Guazzaroni
Jul 8, 2019·World Journal of Microbiology & Biotechnology·Ryosuke MitsuiHiroyasu Ogino
Aug 11, 2019·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Lucas Ferreira RibeiroMaría-Eugenia Guazzaroni
Mar 9, 2019·JACC. Basic to Translational Science·David M GermanSanjiv Kaul
Jan 26, 2021·Nanobiomedicine·Jananee Padayachee, Moganavelli Singh
Apr 28, 2021·Current Protocols·Kornel LabunEivind Valen
Aug 9, 2021·Journal of Controlled Release : Official Journal of the Controlled Release Society·Bixi SunXiaoshu Gao
Feb 2, 2022·Biomaterials Science·Yi LinUlrich Lächelt

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Methods Mentioned

BETA
deamination
acetylation
PCR
fluorescence-activated cell sorting
FACS
one-hybrid

Software Mentioned

PACE
FACS

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