Structure-specific DNA recombination sites: Design, validation, and machine learning-based refinement.

Autor:	Nivina A; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France.; Paris Descartes, Sorbonne Paris Cité, Paris, France., Grieb MS; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France., Loot C; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France., Bikard D; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France., Cury J; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France.; Paris Descartes, Sorbonne Paris Cité, Paris, France., Shehata L; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France., Bernardes J; Laboratoire de Biologie Computationnelle et Quantitative, Sorbonne Universités, CNRS UMR 7238,75005 Paris, France., Mazel D; Unité Plasticité du Génome Bactérien, Institut Pasteur, 75724 Paris, France.; CNRS UMR 3525, 75724 Paris, France.
Jazyk:	angličtina
Zdroj:	Science advances [Sci Adv] 2020 Jul 24; Vol. 6 (30), pp. eaay2922. Date of Electronic Publication: 2020 Jul 24 (Print Publication: 2020).
DOI:	10.1126/sciadv.aay2922
Abstrakt:	Recombination systems are widely used as bioengineering tools, but their sites have to be highly similar to a consensus sequence or to each other. To develop a recombination system free of these constraints, we turned toward attC sites from the bacterial integron system: single-stranded DNA hairpins specifically recombined by the integrase. Here, we present an algorithm that generates synthetic attC sites with conserved structural features and minimal sequence-level constraints. We demonstrate that all generated sites are functional, their recombination efficiency can reach 60%, and they can be embedded into protein coding sequences. To improve recombination of less efficient sites, we applied large-scale mutagenesis and library enrichment coupled to next-generation sequencing and machine learning. Our results validated the efficiency of this approach and allowed us to refine synthetic attC design principles. They can be embedded into virtually any sequence and constitute a unique example of a structure-specific DNA recombination system. (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)
Databáze:	MEDLINE
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