Reference: Sharma J, et al. (2025) Design principle of successful genome editing applications using CRISPR-based toolkits. J Appl Genet

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Abstract


Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins are the most promising toolkit of synthetic biology for genetic engineering applications across species. Essentially, the Type II CRISPR system, featuring Cas9 nuclease from Streptococcus pyogenes complexed with sgRNA, introduces targeted DNA cleavage, enabling modifications with exceptional precision. This technology can be utilized for not only editing but also modulating gene expressions, thereby finding widespread utility in various biotechnological applications. Here we discuss strategies to construct a consolidated platform aiming at developing a CRISPR-based gene editing system in microbial hosts such as yeast. Employing the well-known gene editing enzymes, i.e., Cpf1 and dCas9, two independent strategies to develop a one-pot plasmid system have been proposed. Furthermore, approaches to reduce off-target cleavages introduced by non-specific targeting of CRISPR complex have been discussed. Finally, an overarching discussion on advanced strategies to design robust CRISPR components is provided for streamlining future genome editing applications.

Reference Type
Journal Article | Review
Authors
Sharma J, Biswas R, Khare P
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Gene Ontology Annotations


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Gene/Complex Qualifier Gene Ontology Term Aspect Annotation Extension Evidence Method Source Assigned On Reference

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Gene Phenotype Experiment Type Mutant Information Strain Background Chemical Details Reference

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Gene Disease Ontology Term Qualifier Evidence Method Source Assigned On Reference

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Post-translational Modifications


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Interactor Interactor Allele Assay Annotation Action Phenotype SGA score P-value Source Reference

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Interactor Interactor Assay Annotation Action Modification Source Reference

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Gene Species Gene ID Strain background Direction Details Source Reference