A Cas3-base editing tool for targetable in vivo mutagenesis
Abstract The generation of genetic diversity via mutagenesis is routinely used for protein engineering and pathway optimization. Current technologies for random mutagenesis often target either the whole genome or relatively narrow windows. To bridge this gap, we developed CoMuTER (Confined Mutagenes...
| Main Authors: | , , , , , , |
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| Format: | Article |
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Nature Portfolio
2023-06-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-023-39087-z |
| _version_ | 1797806639135326208 |
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| author | Anna Zimmermann Julian E. Prieto-Vivas Charlotte Cautereels Anton Gorkovskiy Jan Steensels Yves Van de Peer Kevin J. Verstrepen |
| author_facet | Anna Zimmermann Julian E. Prieto-Vivas Charlotte Cautereels Anton Gorkovskiy Jan Steensels Yves Van de Peer Kevin J. Verstrepen |
| author_sort | Anna Zimmermann |
| collection | DOAJ |
| description | Abstract The generation of genetic diversity via mutagenesis is routinely used for protein engineering and pathway optimization. Current technologies for random mutagenesis often target either the whole genome or relatively narrow windows. To bridge this gap, we developed CoMuTER (Confined Mutagenesis using a Type I-E CRISPR-Cas system), a tool that allows inducible and targetable, in vivo mutagenesis of genomic loci of up to 55 kilobases. CoMuTER employs the targetable helicase Cas3, signature enzyme of the class 1 type I-E CRISPR-Cas system, fused to a cytidine deaminase to unwind and mutate large stretches of DNA at once, including complete metabolic pathways. The tool increases the number of mutations in the target region 350-fold compared to the rest of the genome, with an average of 0.3 mutations per kilobase. We demonstrate the suitability of CoMuTER for pathway optimization by doubling the production of lycopene in Saccharomyces cerevisiae after a single round of mutagenesis. |
| first_indexed | 2024-03-13T06:10:19Z |
| format | Article |
| id | doaj.art-4ba11ca207744a24851d08085ba157b2 |
| institution | Directory Open Access Journal |
| issn | 2041-1723 |
| language | English |
| last_indexed | 2024-03-13T06:10:19Z |
| publishDate | 2023-06-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj.art-4ba11ca207744a24851d08085ba157b22023-06-11T11:18:21ZengNature PortfolioNature Communications2041-17232023-06-0114111610.1038/s41467-023-39087-zA Cas3-base editing tool for targetable in vivo mutagenesisAnna Zimmermann0Julian E. Prieto-Vivas1Charlotte Cautereels2Anton Gorkovskiy3Jan Steensels4Yves Van de Peer5Kevin J. Verstrepen6VIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyVIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyVIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyVIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyVIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyDepartment of Plant Biotechnology and Bioinformatics, Ghent UniversityVIB Laboratory for Systems Biology, VIB-KU Leuven Center for MicrobiologyAbstract The generation of genetic diversity via mutagenesis is routinely used for protein engineering and pathway optimization. Current technologies for random mutagenesis often target either the whole genome or relatively narrow windows. To bridge this gap, we developed CoMuTER (Confined Mutagenesis using a Type I-E CRISPR-Cas system), a tool that allows inducible and targetable, in vivo mutagenesis of genomic loci of up to 55 kilobases. CoMuTER employs the targetable helicase Cas3, signature enzyme of the class 1 type I-E CRISPR-Cas system, fused to a cytidine deaminase to unwind and mutate large stretches of DNA at once, including complete metabolic pathways. The tool increases the number of mutations in the target region 350-fold compared to the rest of the genome, with an average of 0.3 mutations per kilobase. We demonstrate the suitability of CoMuTER for pathway optimization by doubling the production of lycopene in Saccharomyces cerevisiae after a single round of mutagenesis.https://doi.org/10.1038/s41467-023-39087-z |
| spellingShingle | Anna Zimmermann Julian E. Prieto-Vivas Charlotte Cautereels Anton Gorkovskiy Jan Steensels Yves Van de Peer Kevin J. Verstrepen A Cas3-base editing tool for targetable in vivo mutagenesis Nature Communications |
| title | A Cas3-base editing tool for targetable in vivo mutagenesis |
| title_full | A Cas3-base editing tool for targetable in vivo mutagenesis |
| title_fullStr | A Cas3-base editing tool for targetable in vivo mutagenesis |
| title_full_unstemmed | A Cas3-base editing tool for targetable in vivo mutagenesis |
| title_short | A Cas3-base editing tool for targetable in vivo mutagenesis |
| title_sort | cas3 base editing tool for targetable in vivo mutagenesis |
| url | https://doi.org/10.1038/s41467-023-39087-z |
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