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dc.contributor.authorCampelo, Ana B.-
dc.contributor.authorRoces, Clara-
dc.contributor.authorMohedano Bonillo, Mari Luz-
dc.contributor.authorLópez, Paloma-
dc.contributor.authorRodríguez González, Ana-
dc.contributor.authorMartínez Fernández, Beatriz-
dc.identifier.citationMicrobial Cell Factories 13: 77 (2014)es_ES
dc.description.abstract[Background] Lactococcus lactis is widely used as a dairy starter and has been extensively studied. Based on the acquired knowledge on its physiology and metabolism, new applications have been envisaged and there is an increasing interest of using L. lactis as a cell factory. Plasmids constitute the main toolbox for L. lactis genetic engineering and most rely on antibiotic resistant markers for plasmid selection and maintenance. In this work, we have assessed the ability of the bacteriocin Lactococcin 972 (Lcn972) gene cluster to behave as a food-grade post-segregational killing system to stabilize recombinant plasmids in L. lactis in the absence of antibiotics. Lcn972 is a non-lantibiotic bacteriocin encoded by the 11-kbp plasmid pBL1 with a potent antimicrobial activity against Lactococcus.es_ES
dc.description.abstract[Results] Attempts to clone the full lcn972 operon with its own promoter (P972), the structural gene lcn972 and the immunity genes orf2-orf3 in the unstable plasmid pIL252 failed and only plasmids with a mutated promoter were recovered. Alternatively, cloning under other constitutive promoters was approached and achieved, but bacteriocin production levels were lower than those provided by pBL1. Segregational stability studies revealed that the recombinant plasmids that yielded high bacteriocin titers were maintained for at least 200 generations without antibiotic selection. In the case of expression vectors such as pTRL1, the Lcn972 gene cluster also contributed to plasmid maintenance without compromising the production of the fluorescent mCherry protein. Furthermore, unstable Lcn972 recombinant plasmids became integrated into the chromosome through the activity of insertion sequences, supporting the notion that Lcn972 does apply a strong selective pressure against susceptible cells. Despite of it, the Lcn972 gene cluster was not enough to avoid the use of antibiotics to select plasmid-bearing cells right after transformation.es_ES
dc.description.abstract[Conclusions] Inserting the Lcn972 cluster into segregational unstable plasmids prevents their lost by segregation and probable could be applied as an alternative to the use of antibiotics to support safer and more sustainable biotechnological applications of genetically engineered L. lactis.es_ES
dc.description.sponsorshipThis work has been funded by grant BIO2010-17414 (Ministerio de Ciencia e Innovación, Spain). Work of PL and MLM was supported by grant AGL2012-40084-C03-01 (Ministerio de Economía y Competitividad, Spain). We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI). C.R. was a recipient of a predoctoral JAE-CSIC fellowship. We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI)es_ES
dc.publisherBioMed Centrales_ES
dc.relation.isversionofPublisher's versiones_ES
dc.subjectPlasmid segregationes_ES
dc.subjectPost-segregational killinges_ES
dc.subjectInsertion sequenceses_ES
dc.subjectLactococcus lactises_ES
dc.titleA bacteriocin gene cluster able to enhance plasmid maintenance in Lactococcus lactises_ES
dc.description.peerreviewedPeer reviewedes_ES
dc.contributor.funderMinisterio de Ciencia e Innovación (España)-
dc.contributor.funderMinisterio de Economía y Competitividad (España)-
dc.contributor.funderCSIC - Unidad de Recursos de Información Científica para la Investigación (URICI)-
dc.contributor.funderEuropean Commission-
dc.contributor.funderConsejo Superior de Investigaciones Científicas (España)-
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