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dc.contributor.author
Volke, Daniel C.  
dc.contributor.author
Martino, Román Alejandro  
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Kozaeva, Ekaterina  
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Smania, Andrea  
dc.contributor.author
Nikel, Pablo I.  
dc.date.available
2024-01-11T12:16:03Z  
dc.date.issued
2022-05  
dc.identifier.citation
Volke, Daniel C.; Martino, Román Alejandro; Kozaeva, Ekaterina; Smania, Andrea; Nikel, Pablo I.; Modular (de)construction of complex bacterial phenotypes by CRISPR/nCas9-assisted, multiplex cytidine base-editing; Springer; Nature Communications; 13; 1; 5-2022; 1-14  
dc.identifier.issn
2041-1723  
dc.identifier.uri
http://hdl.handle.net/11336/223304  
dc.description.abstract
CRISPR/Cas technologies constitute a powerful tool for genome engineering, yet their use in non-traditional bacteria depends on host factors or exogenous recombinases, which limits both efficiency and throughput. Here we mitigate these practical constraints by developing a widely-applicable genome engineering toolset for Gram-negative bacteria. The challenge is addressed by tailoring a CRISPR base editor that enables single-nucleotide resolution manipulations (C·G → T·A) with >90% efficiency. Furthermore, incorporating Cas6-mediated processing of guide RNAs in a streamlined protocol for plasmid assembly supports multiplex base editing with >85% efficiency. The toolset is adopted to construct and deconstruct complex phenotypes in the soil bacterium Pseudomonas putida. Single-step engineering of an aromatic-compound production phenotype and multi-step deconstruction of the intricate redox metabolism illustrate the versatility of multiplex base editing afforded by our toolbox. Hence, this approach overcomes typical limitations of previous technologies and empowers engineering programs in Gram-negative bacteria that were out of reach thus far.  
dc.format
application/pdf  
dc.language.iso
eng  
dc.publisher
Springer  
dc.rights
info:eu-repo/semantics/openAccess  
dc.rights.uri
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/  
dc.subject
CRISPR/Cas9  
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Base editors  
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Pseudomonas spp  
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Multiplex genome editing  
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Bioquímica y Biología Molecular  
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Ciencias Biológicas  
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CIENCIAS NATURALES Y EXACTAS  
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Biología Celular, Microbiología  
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Ciencias Biológicas  
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CIENCIAS NATURALES Y EXACTAS  
dc.title
Modular (de)construction of complex bacterial phenotypes by CRISPR/nCas9-assisted, multiplex cytidine base-editing  
dc.type
info:eu-repo/semantics/article  
dc.type
info:ar-repo/semantics/artículo  
dc.type
info:eu-repo/semantics/publishedVersion  
dc.date.updated
2024-01-11T09:55:07Z  
dc.journal.volume
13  
dc.journal.number
1  
dc.journal.pagination
1-14  
dc.journal.pais
Reino Unido  
dc.description.fil
Fil: Volke, Daniel C.. Technical University of Denmark; Dinamarca  
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Fil: Martino, Román Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica; Argentina  
dc.description.fil
Fil: Kozaeva, Ekaterina. Technical University of Denmark; Dinamarca  
dc.description.fil
Fil: Smania, Andrea. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentina  
dc.description.fil
Fil: Nikel, Pablo I.. Technical University of Denmark; Dinamarca  
dc.journal.title
Nature Communications  
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/url/https://www.nature.com/articles/s41467-022-30780-z#citeas  
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info:eu-repo/semantics/altIdentifier/doi/https://doi.org/10.1038/s41467-022-30780-z