An Episomal CRISPR/Cas12a System for Mediating Efficient Gene Editing
Abstract
:1. Introduction
2. Materials and Methods
2.1. Design and Construction of Episomal CRISPR/Cas12a
2.2. Cell Culture and Maintenance
2.3. Genome Editing of HEK-293T with COE System
2.4. Restriction Fragment Length Polymorphism (RFLP) Analysis
2.5. Genome Editing of iPSCs with COE System
2.6. RNA Isolation and Quantitative Polymerase Chain Reaction (qPCR)
2.7. Cardiac Differentiation
2.8. Immunofluorescence Staining
2.9. Residue of Plasmid Components and Off-Target Analysis
2.10. Statistical Analysis
3. Results
3.1. Establishment of the Episomal CRISPR/Cas12a System
3.2. The COE System Significantly Promotes Gene Editing in HEK-293T
3.3. The COE System Efficient for Gene Deletion in Human iPSCs
3.4. The Edited iPSCs Were Directed to Differentiate into Cardiomyocytes
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Duan, N.; Tang, S.; Zeng, B.; Hu, Z.; Hu, Q.; Wu, L.; Zhou, M.; Liang, D. An Episomal CRISPR/Cas12a System for Mediating Efficient Gene Editing. Life 2021, 11, 1262. https://doi.org/10.3390/life11111262
Duan N, Tang S, Zeng B, Hu Z, Hu Q, Wu L, Zhou M, Liang D. An Episomal CRISPR/Cas12a System for Mediating Efficient Gene Editing. Life. 2021; 11(11):1262. https://doi.org/10.3390/life11111262
Chicago/Turabian StyleDuan, Nannan, Shuqing Tang, Baitao Zeng, Zhiqing Hu, Qian Hu, Lingqian Wu, Miaojin Zhou, and Desheng Liang. 2021. "An Episomal CRISPR/Cas12a System for Mediating Efficient Gene Editing" Life 11, no. 11: 1262. https://doi.org/10.3390/life11111262