60993 20190709135532.0 doi 10.3389/fmicb.2017.00315 sideral 98571 ART-2017-98571 eng (orcid)0000-0003-1027-7583 Otal, I. Universidad de Zaragoza Detection of a putative TetR-like gene related to Mycobacterium bovis BCG growth in cholesterol using a gfp-transposon mutagenesis system 2017 Access copy available to the general public Unrestricted In vitro transposition is a powerful genetic tool for identifying mycobacterial virulence genes and studying virulence factors in relation to the host. Transposon shuttle mutagenesis is a method for constructing stable insertions in the genome of different microorganisms including mycobacteria. Using an IS1096 derivative, we have constructed the Tngfp, a transposon containing a promoterless green fluorescent protein (gfp) gene. This transposon was able to transpose randomly in Mycobacterium bovis BCG. Bacteria with a single copy of the gfp gene per chromosome from an M. bovis BCG::Tngfp library were analyzed and cells exhibiting high levels of fluorescence were detected by flow cytometry. Application of this approach allowed for the selection of a mutant, BCG_2177c::Tngfp (BCG-Tn), on the basis of high level of long-standing fluorescence at stationary phase. This BCG-Tn mutant showed some particular phenotypic features compared to the wild type strain, mainly during stationary phase, when cholesterol was used as a sole carbon source, thus supporting the relationships of the targeted gene with the regulation of cholesterol metabolism in this bacteria. This approach showed that Tngfp is a potentially useful tool for studying the involvement of the targeted loci in metabolic pathways of mycobacteria. info:eu-repo/grantAgreement/ES/DGA/B25 info:eu-repo/grantAgreement/ES/MEC/SAB2010-0102 info:eu-repo/grantAgreement/ES/MINECO/BIO2014-5258P info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 4.019 2017 MICROBIOLOGY 31 / 125 = 0.248 2017 Q1 T1 1.699 2017 Microbiology (medical) 2017 Q1 Microbiology 2017 Q1 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Pérez-Herrán, E. Garcia-Morales, L. Menéndez, M. C. Gonzalez-y-Merchand, J. A. (orcid)0000-0003-2993-5478 Martín, C. Universidad de Zaragoza García, M. J. 1008 630 Universidad de Zaragoza Dpto. Microb.Med.Pr.,Sal.Públ. Área Microbiología 8 (2017), 315 [13 pp.] Front. microbiol. FRONTIERS IN MICROBIOLOGY 1664-302X 2114528 http://zaguan.unizar.es/record/60993/files/texto_completo.pdf Versión publicada 22586 http://zaguan.unizar.es/record/60993/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:60993 articulos driver 2019-07-09-12:03:01 ARTICLE