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I-SceI-mediated plasmid deletion and intra-molecular recombination in Spiroplasma citri

hal.structure.identifierGénomique, développement et pouvoir pathogène [GD2P]
dc.contributor.authorBRETON, Marc
hal.structure.identifierGénomique, développement et pouvoir pathogène [GD2P]
dc.contributor.authorDURET, Sybille
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorDANET, Jean-Luc
hal.structure.identifierGénomique, développement et pouvoir pathogène [GD2P]
dc.contributor.authorBEVEN, Laure
hal.structure.identifierGénomique, développement et pouvoir pathogène [GD2P]
dc.contributor.authorDUBRANA, Marie-Pierre
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorRENAUDIN, Joël
dc.date.issued2011
dc.identifier.issn0167-7012
dc.description.abstractEnS. citri wild-type strain GII3 carries six plasmids (pSci1 to -6) that are thought to encode determinants involved in the transmission of the spiroplasma by its leafhopper vector. In this study we report the use of meganuclease I-SceI for plasmid deletion in S. citri. Plasmids pSci1NT-I and pSci6PT-I, pSci1 and pSci6 derivatives that contain the tetM selection marker and a unique I-SceI recognition site were first introduced into S. citri strains 44 (having no plasmid) and GII3 (carrying pSci1–6), respectively. Due to incompatibility of homologous replication regions, propagation of the S. citri GII3 transformant in selective medium resulted in the replacement of the natural pSci6 by pSci6PT-I. The spiroplasmal transformants were further transformed by an oriC plasmid carrying the I-SceI gene under the control of the spiralin gene promoter. In the S. citri 44 transformant, expression of I-SceI resulted in rapid loss of pSciNT-I showing that expression of I-SceI can be used as a counter-selection tool in spiroplasmas. In the case of the S. citri GII3 transformant carrying pSci6PT-I, expression of I-SceI resulted in the deletion of plasmid fragments comprising the I-SceI site and the tetM marker. Delineating the I-SceI generated deletions proved they had occurred though recombination between homologous sequences. To our knowledge this is the first report of I-SceI mediated intra-molecular recombination in mollicutes.
dc.language.isoen
dc.publisherElsevier
dc.subjectMOLLICUTES
dc.subjectBACTERIOLOGIE
dc.subjectGENETIQUE
dc.subject.enSPIROPLASMA CITRI
dc.subject.enPLASMIDS
dc.subject.enENDONUCLEASE I-SCEL
dc.subject.enCOUNTER-SELECTION
dc.subject.enHOMOLOGOUS RECOMBINATION
dc.subject.enCONTRE-SELECTION
dc.title.enI-SceI-mediated plasmid deletion and intra-molecular recombination in Spiroplasma citri
dc.typeArticle de revue
dc.identifier.doi10.1016/j.mimet.2010.11.020
dc.subject.halSciences du Vivant [q-bio]/Microbiologie et Parasitologie
bordeaux.journalJournal of Microbiological Methods
bordeaux.page216-222
bordeaux.volume84
bordeaux.issue2
bordeaux.peerReviewedoui
hal.identifierhal-02649350
hal.version1
hal.popularnon
hal.audienceInternationale
hal.origin.linkhttps://hal.archives-ouvertes.fr//hal-02649350v1
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