PUBLICATION
Efficient and risk-reduced genome editing using double nicks enhanced by bacterial recombination factors in multiple species
- Authors
- He, X., Chen, W., Liu, Z., Yu, G., Chen, Y., Cai, Y.J., Sun, L., Xu, W., Zhong, L., Gao, C., Chen, J., Zhang, M., Yang, S., Yao, Y., Zhang, Z., Ma, F., Zhang, C.C., Lu, H.P., Yu, B., Cheng, T.L., Qiu, J., Sheng, Q., Zhou, H.M., Lv, Z.R., Yan, J., Zhou, Y., Qiu, Z., Cui, Z., Zhang, X., Meng, A., Sun, Q., Yang, Y.
- ID
- ZDB-PUB-200403-223
- Date
- 2020
- Source
- Nucleic acids research 48(10): e57 (Journal)
- Registered Authors
- Meng, Anming
- Keywords
- none
- MeSH Terms
-
- Animals
- Bacterial Proteins/metabolism*
- DNA Breaks, Double-Stranded
- DNA-Binding Proteins/metabolism
- Female
- Gene Editing/methods*
- Gene Knock-In Techniques
- Genomics
- Homologous Recombination
- Humans
- INDEL Mutation
- Macaca fascicularis
- Mice
- Rats, Sprague-Dawley
- Rec A Recombinases/metabolism
- Zebrafish/genetics
- PubMed
- 32232370 Full text @ Nucleic Acids Res.
Citation
He, X., Chen, W., Liu, Z., Yu, G., Chen, Y., Cai, Y.J., Sun, L., Xu, W., Zhong, L., Gao, C., Chen, J., Zhang, M., Yang, S., Yao, Y., Zhang, Z., Ma, F., Zhang, C.C., Lu, H.P., Yu, B., Cheng, T.L., Qiu, J., Sheng, Q., Zhou, H.M., Lv, Z.R., Yan, J., Zhou, Y., Qiu, Z., Cui, Z., Zhang, X., Meng, A., Sun, Q., Yang, Y. (2020) Efficient and risk-reduced genome editing using double nicks enhanced by bacterial recombination factors in multiple species. Nucleic acids research. 48(10):e57.
Abstract
Site-specific DNA double-strand breaks have been used to generate knock-in through the homology-dependent or -independent pathway. However, low efficiency and accompanying negative impacts such as undesirable indels or tumorigenic potential remain problematic. In this study, we present an enhanced reduced-risk genome editing strategy we named as NEO, which used either site-specific trans or cis double-nicking facilitated by four bacterial recombination factors (RecOFAR). In comparison to currently available approaches, NEO achieved higher knock-in (KI) germline transmission frequency (improving from zero to up to 10% efficiency with an average of 5-fold improvement for 8 loci) and 'cleaner' knock-in of long DNA fragments (up to 5.5 kb) into a variety of genome regions in zebrafish, mice and rats. Furthermore, NEO yielded up to 50% knock-in in monkey embryos and 20% relative integration efficiency in non-dividing primary human peripheral blood lymphocytes (hPBLCs). Remarkably, both on-target and off-target indels were effectively suppressed by NEO. NEO may also be used to introduce low-risk unrestricted point mutations effectively and precisely. Therefore, by balancing efficiency with safety and quality, the NEO method reported here shows substantial potential and improves the in vivo gene-editing strategies that have recently been developed.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping