Detection of a putative TetR-like gene related to Mycobacterium bovis BCG growth in cholesterol using a gfp-transposon mutagenesis system

Otal, I. (Universidad de Zaragoza) ; Pérez-Herrán, E. ; Garcia-Morales, L. ; Menéndez, M. C. ; Gonzalez-y-Merchand, J. A. ; Martín, C. (Universidad de Zaragoza) ; García, M. J.

Resumen: 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.

Idioma: Inglés

DOI: 10.3389/fmicb.2017.00315

Año: 2017

Publicado en: FRONTIERS IN MICROBIOLOGY 8 (2017), 315 [13 pp.]
ISSN: 1664-302X

Factor impacto JCR: 4.019 (2017)

Categ. JCR: MICROBIOLOGY rank: 31 / 125 = 0.248 (2017) - Q1 - T1

Factor impacto SCIMAGO: 1.699 - Microbiology (medical) (Q1) - Microbiology (Q1)

Financiación: info:eu-repo/grantAgreement/ES/DGA/B25
Financiación: info:eu-repo/grantAgreement/ES/MEC/SAB2010-0102
Financiación: info:eu-repo/grantAgreement/ES/MINECO/BIO2014-5258P

Tipo y forma: Article (Published version)

Área (Departamento): Área Microbiología (Dpto. Microb.Med.Pr.,Sal.Públ.)