Abstract
Background: RNA-guided endonuclease as a versatile genome editing technology opened new windows in various fields of biology. The simplicity of this revolutionary technique provides a promising future for its application in a broad range of approaches from functional annotation of genes to diseases, to genetic manipulation and gene therapy. Besides the site-specific activity of Cas9 endonuclease, the unintended cleavage known as off-target effect is still a major challenge for this genome editing technique.
Methods: Various strategies have been developed to resolve this bottleneck including development of new softwares for designing optimized guide RNA (gRNA), engineering Cas9 enzyme, improvement in off-target detection assays, etc.
Results: This review dedicated to discuss on methods that have been used for optimizing Cas9, specificity with the aim of improving this technology for therapeutic applications.
Conclusion: In addition, the applications and novel breakthroughs in the field of CRISPR technology will be described.
Keywords: CRISPR-Cas systems, guide RNA (gRNA), gene editing, mutation, off-target effect, single-strand specific DNA and RNA endonucleases.
Graphical Abstract
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