title: Construction and evaluation of a chromosomal expression platform (CEP) for ectopic, maltose-driven gene expression in Streptococcus pneumoniae.
abstract: In this paper, the construction and evaluation of a chromosomal expression platform (CEP), which allows controlled gene expression following ectopic integration into the chromosome of Streptococcus pneumoniae, is described. CEP is based on the well-studied maltosaccharide-inducible system. To facilitate integration at CEP, a plasmid, pCEP, capable of replication in Escherichia coli, but not in S. pneumoniae, was assembled. This plasmid contains an expression/selection cassette flanked on each side by more than 2 kb of pneumococcal DNA. The cassette comprises a maltose-inducible promoter, P(M), separated from a kanamycin-resistance gene by NcoI and BamHI cloning sites. Clones harbouring the gene of interest integrated at CEP under the control of P(M) can be obtained through direct transformation of an S. pneumoniae recipient with ligation products between that gene and NcoI/BamHI-digested pCEP DNA, followed by selection for kanamycin-resistant transformants.
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Guiral
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Hénard
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Maria-Halima
authors.initials: MH
authors.lastname: Laaberki
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Chantal
authors.initials: C
authors.lastname: Granadel
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Marc
authors.initials: M
authors.lastname: Prudhomme
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Bernard
authors.initials: B
authors.lastname: Martin
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100 CNRS-Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex 9, France.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Claverys
mesh_terms: D001431:Bacteriological Techniques; D002876:Chromosomes, Bacterial / Q000502:physiology*; D003433:Crosses, Genetic; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D008320:Maltose / Q000502:physiology*; D013296:Streptococcus pneumoniae / Q000235:genetics* / Q000378:metabolism
chemical_list: D008320:Maltose