Mismatch intolerance of 5′-truncated sgrnas in crispr/cas9 enables efficient microbial single-base genome editingopen access
- Authors
- Lee, Ho Joung; Kim, Hyun Ju; Lee, Sang Jun
- Issue Date
- Jun-2021
- Publisher
- MDPI AG
- Keywords
- 5′-truncated sgRNA; Cas9; CRISPR; Microbial genome editing; Single base
- Citation
- International Journal of Molecular Sciences, v.22, no.12
- Journal Title
- International Journal of Molecular Sciences
- Volume
- 22
- Number
- 12
- URI
- https://scholarworks.bwise.kr/cau/handle/2019.sw.cau/48929
- DOI
- 10.3390/ijms22126457
- ISSN
- 1661-6596
1422-0067
- Abstract
- The CRISPR/Cas9 system has recently emerged as a useful gene-specific editing tool. However, this approach occasionally results in the digestion of both the DNA target and similar DNA sequences due to mismatch tolerance, which remains a significant drawback of current genome editing technologies. However, our study determined that even single-base mismatches between the target DNA and 5′-truncated sgRNAs inhibited target recognition. These results suggest that a 5′-truncated sgRNA/Cas9 complex could be used to negatively select single-base-edited targets in microbial genomes. Moreover, we demonstrated that the 5′-truncated sgRNA method can be used for simple and effective single-base editing, as it enables the modification of individual bases in the DNA target, near and far from the 5′ end of truncated sgRNAs. Further, 5′-truncated sgRNAs also allowed for efficient single-base editing when using an engineered Cas9 nuclease with an expanded protospacer adjacent motif (PAM; 5′-NG), which may enable whole-genome single-base editing. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.
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Collections - College of Biotechnology & Natural Resource > Department of Systems Biotechnology > 1. Journal Articles
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