Engineering of CRISPR-Cas12b for human genome editing.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 30670702.
- Also identified by DOI 10.1038/s41467-018-08224-4 and PMC identifier 6342934.
- Licence recorded as CC BY.
- The licence permits redistribution, so the abstract is shown in full and the full text is available from the publisher.
Abstract
The type-V CRISPR effector Cas12b (formerly known as C2c1) has been challenging to develop for genome editing in human cells, at least in part due to the high temperature requirement of the characterized family members. Here we explore the diversity of the Cas12b family and identify a promising candidate for human gene editing from Bacillus hisashii, BhCas12b. However, at 37 °C, wild-type BhCas12b preferentially nicks the non-target DNA strand instead of forming a double strand break, leading to lower editing efficiency. Using a combination of approaches, we identify gain-of-function mutations for BhCas12b that overcome this limitation. Mutant BhCas12b facilitates robust genome editing in human cell lines and ex vivo in primary human T cells, and exhibits greater specificity compared to S. pyogenes Cas9. This work establishes a third RNA-guided nuclease platform, in addition to Cas9 and Cpf1/Cas12a, for genome editing in human cells.
Medical subject headings
- Bacillus
- Bacterial Proteins
- CRISPR-Cas Systems
- Deoxyribonucleases
- Gene Editing
- Genome, Human