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CRISPR/Cas9 in mycoplasma: from natural systems of minimal bacteria towards tools for genome engineering

hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorIPOUTCHA, Thomas
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorTSARMPOPOULOS, Iason
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorGOURGUES, Geraldine
hal.structure.identifierLabri, Univ. Bordeaux, Bordeaux
dc.contributor.authorTHEBAULT, Patricia
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorBLANCHARD, Alain
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorLARTIGUE, Carole
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorSIRAND-PUGNET, Pascal
dc.date.accessioned2024-09-12T02:03:45Z
dc.date.available2024-09-12T02:03:45Z
dc.date.conference2021-06-01
dc.identifier.urihttps://oskar-bordeaux.fr/handle/20.500.12278/201536
dc.description.abstractEnMollicutes are minimal bacteria that have no cell wall and most of them use UGA stop codon to encode Trp. They are considered as the smallest bacteria able to grow in axenic media. Their genomes are AT-rich, with sizes ranging from 580 kbp to 2.2 Mbp. Mollicutes are currently the bacteria with the smallest genome in which CRISPR/Cas systems have been reported. Complete or degraded systems were found in the genome of 21 out of 52 representative Mollicutes species, and most of them belong to the Type II-A and some to the Type II-C. We characterized a representative CRISPR/Cas9 system of mycoplasmas identified in the bird pathogen Mycoplasma gallisepticum strain S6, MgalCas9. In silico analysis and in vivo cleavage assay demonstrated functional activity of MgalCas9 in this strain. In vitro analyses by plasmid library cleavage indicated a 5-positions PAM consensus (NNAAT). This PAM preference was shown clearly different in M. gallisepticum strain CA06 (NNGAD), suggesting a recent intraspecific evolution of MgCas9. Mycoplasma lacking of genetic tools, we developed a genome engineering tool based on MgalCas9 which efficiency was evaluated in a phylogenetically remote species, Mycoplasma capricolum. An all-in-one oriC plasmid harbouring genes encoding MgalCas9 and a sgRNA was built. We demonstrated the functionality of this tool in different mollicutes species including M. capricolum, Mycoplasma mycoides, Mycoplasma pulmonis and M. gallisepticum. Current work aims at developing other tools as Cas9-deaminase fusion protein and additional exogenous recombination systems for the genome engineering of these minimal bacteria.
dc.language.isoen
dc.title.enCRISPR/Cas9 in mycoplasma: from natural systems of minimal bacteria towards tools for genome engineering
dc.typeCommunication dans un congrès
dc.subject.halSciences du Vivant [q-bio]/Microbiologie et Parasitologie
bordeaux.hal.laboratoriesBiologie du Fruit & Pathologie (BFP) - UMR 1332*
bordeaux.institutionUniversité de Bordeaux
bordeaux.institutionINRAE
bordeaux.conference.titleCRISPR conference 2021 (Virtuel)
bordeaux.countryFR
bordeaux.conference.cityParis
bordeaux.peerReviewedoui
hal.identifierhal-04694342
hal.version1
hal.invitednon
hal.proceedingsnon
hal.conference.end2021-06-10
hal.popularnon
hal.audienceInternationale
hal.origin.linkhttps://hal.archives-ouvertes.fr//hal-04694342v1
bordeaux.COinSctx_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.au=IPOUTCHA,%20Thomas&TSARMPOPOULOS,%20Iason&GOURGUES,%20Geraldine&THEBAULT,%20Patricia&BLANCHARD,%20Alain&rft.genre=unknown


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