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Base-Editor-Mediated circRNA Knockout by Targeting Predominantly Back-Splice Sites.
Xu-Kai Ma1, Xiang Gao2,3, Mei Cao1
1Center for Molecular Medicine, Children's Hospital, Fudan University and Shanghai Key Laboratory of Medical Epigenetics, International Laboratory of Medical Epigenetics and Metabolism, Institutes of Biomedical Sciences, Fudan University, Shanghai, China.
Methods in Molecular Biology (Clifton, N.J.)
|February 21, 2024
Summary
This study introduces a novel method for selectively removing circular RNAs (circRNAs) by targeting their unique back-splice sites using base editing technology. This approach enables efficient circRNA knockout (KO) at the genomic DNA level.
Area of Science:
- Molecular Biology
- Genetics
- RNA Biology
Background:
- Circular RNAs (circRNAs) are formed by back-splicing of eukaryotic exons.
- Distinguishing circRNAs from their linear counterparts is challenging due to sequence overlap.
- Existing methods for circRNA manipulation are limited.
Purpose of the Study:
- To present a detailed protocol for circRNA knockout (KO) using base editors.
- To target and disrupt the back-splice sites crucial for circRNA formation.
- To provide a method for specific depletion of circRNAs at the genomic DNA level.
Main Methods:
- Utilizing base editing systems to target specific back-splice sites.
- Designing guide RNAs to direct base editors to circRNA formation junctions.
- Applying the protocol to achieve genomic DNA-level disruption of circRNA biogenesis.
Main Results:
- Demonstrated successful depletion of specific circRNAs via base editing.
- Showcased the efficiency of targeting back-splice sites for circRNA KO.
- Validated the protocol for precise circRNA disruption.
Conclusions:
- Base editing offers an efficient and specific strategy for circRNA knockout.
- The described protocol facilitates targeted circRNA depletion at the genomic DNA level.
- This method advances the study of circRNA function through targeted genetic manipulation.
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