A RAGE based strategy for the genome engineering of the human respiratory pathogen mycoplasma pneumoniae
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- dc.contributor.author García Morales, Luis
- dc.contributor.author Ruiz, Estelle
- dc.contributor.author Gourgues, Géraldine
- dc.contributor.author Rideau, Fabien
- dc.contributor.author Piñero-Lambea, Carlos
- dc.contributor.author Lluch-Senar, Maria 1982-
- dc.contributor.author Blanchard, Alain
- dc.contributor.author Lartigue, Carole
- dc.date.accessioned 2020-11-12T11:43:28Z
- dc.date.available 2020-11-12T11:43:28Z
- dc.date.issued 2020
- dc.description.abstract Genome engineering of microorganisms has become a standard in microbial biotechnologies. Several efficient tools are available for the genetic manipulation of model bacteria such as Escherichia coli and Bacillus subtilis, or the yeast Saccharomyces cerevisiae. Difficulties arise when transferring these tools to nonmodel organisms. Synthetic biology strategies relying on genome transplantation (GT) aim at using yeast cells for engineering bacterial genomes cloned as artificial chromosomes. However, these strategies remain unsuccessful for many bacteria, including Mycoplasma pneumoniae (MPN), a human pathogen infecting the respiratory tract that has been extensively studied as a model for systems biology of simple unicellular organisms. Here, we have designed a novel strategy for genome engineering based on the recombinase-assisted genomic engineering (RAGE) technology for editing the MPN genome. Using this strategy, we have introduced a 15 kbp fragment at a specific locus of the MPN genome and replaced 38 kbp from its genome by engineered versions modified either in yeast or in E. coli. A strain harboring a synthetic version of this fragment cleared of 13 nonessential genes could also be built and propagated in vitro. These strains were depleted of known virulence factors aiming at creating an avirulent chassis for SynBio applications. Such a chassis and technology are a step forward to build vaccines or deliver therapeutic compounds in the lungs to prevent or cure respiratory diseases in humans.
- dc.description.sponsorship The authors thank Clara Blanchard for English proofreading. We also thank Dr. Don L. Court for providing biological material and advice. We acknowledge the financial support by the European Union’s Horizon 2020 research and innovation program under Grant Agreement [No. 634942]; and the European MiniCell project selected by ANR, in the frame of the ERASynBio second Joint Call for Transnational Research Projects [No. ANR-15-SYNB-0001-04].
- dc.format.mimetype application/pdf
- dc.identifier.citation Garcia-Morales L, Ruiz E, Gourgues G, Rideau F, Piñero-Lambea C, Lluch-Senar M, Blanchard A, Lartigue C. A RAGE based strategy for the genome engineering of the human respiratory pathogen mycoplasma pneumoniae. ACS Synth Biol. 2020; 9(10):2737-48. DOI: 10.1021/acssynbio.0c00263
- dc.identifier.doi http://dx.doi.org/10.1021/acssynbio.0c00263
- dc.identifier.issn 2161-5063
- dc.identifier.uri http://hdl.handle.net/10230/45739
- dc.language.iso eng
- dc.publisher American Chemical Society (ACS)
- dc.relation.ispartof ACS Synth Biol. 2020; 9(10):2737-48
- dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/634942
- dc.rights © 2020 American Chemical Society. This is an open access article published under an ACS AuthorChoice License (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html), which permitscopying and redistribution of the article or any adaptations for non-commercial purposes.
- dc.rights.accessRights info:eu-repo/semantics/openAccess
- dc.rights.uri http://pubs.acs.org/page/policy/authorchoice_termsofuse.html
- dc.subject.keyword Mycoplasma pneumoniae (MPN)
- dc.subject.keyword Saccharomyces cerevisiae
- dc.subject.keyword Genome editing
- dc.subject.keyword Phage recombinases
- dc.subject.keyword Recombinase-assisted genomic engineering (RAGE)
- dc.subject.keyword Transformation-associated recombination-cloning (TAR-cloning)
- dc.title A RAGE based strategy for the genome engineering of the human respiratory pathogen mycoplasma pneumoniae
- dc.type info:eu-repo/semantics/article
- dc.type.version info:eu-repo/semantics/publishedVersion