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Characterization of a thermostable Cas12a ortholog
- Source :
- Cell Insight, Vol 2, Iss 6, Pp 100126- (2023)
- Publication Year :
- 2023
- Publisher :
- Elsevier, 2023.
-
Abstract
- CRISPR-Cas12a has been used for genome editing and molecular diagnosis. The well-studied Cas12a orthologs have a T-rich PAM and are usually categorized as non-thermally stable enzymes. Here, we identified a new Cas12a ortholog from Clostridium thermobutyricum, which survives at 60 °C. This Cas12a ortholog is named as CtCas12a and exhibits low sequence similarity to the known Cas12a family members. CtCas12a is active in a wide temperature range from 17 to 77 °C. Moreover, this ortholog has a relaxed PAM of YYV (Y=C or T, V = A or C or G). We optimized the conditions for trans-cleavage and enabled its detection of nucleic acids. CtCas12a executed genome editing in human cells and generated up to 26% indel formation in the EGFP locus. With the ability to be active at high temperatures as well as having a relaxed PAM sequence, CtCas12a holds potential to be further engineered for pathogen detection and editing a wide range of genomic sequences.
- Subjects :
- Biology (General)
QH301-705.5
Medicine (General)
R5-920
Subjects
Details
- Language :
- English
- ISSN :
- 27728927
- Volume :
- 2
- Issue :
- 6
- Database :
- Directory of Open Access Journals
- Journal :
- Cell Insight
- Publication Type :
- Academic Journal
- Accession number :
- edsdoj.91490d19e4144362836042d735bc527f
- Document Type :
- article
- Full Text :
- https://doi.org/10.1016/j.cellin.2023.100126