title: Improved electrotransformation and decreased antibiotic resistance of the cystic fibrosis pathogen Burkholderia cenocepacia strain J2315.
abstract: The bacterium Burkholderia cenocepacia is pathogenic for sufferers from cystic fibrosis (CF) and certain immunocompromised conditions. The B. cenocepacia strain most frequently isolated from CF patients, and which serves as the reference for CF epidemiology, is J2315. The J2315 genome is split into three chromosomes and one plasmid. The strain was sequenced several years ago, and its annotation has been released recently. This information should allow genetic experimentation with J2315, but two major impediments appear: the poor potential of J2315 to act as a recipient in transformation and conjugation and the high level of resistance it mounts to nearly all antibiotics. Here, we describe modifications to the standard electroporation procedure that allow routine transformation of J2315 by DNA. In addition, we show that deletion of an efflux pump gene and addition of spermine to the medium enhance the sensitivity of J2315 to certain commonly used antibiotics and so allow a wider range of antibiotic resistance genes to be used for selection.
authors.affiliation: Laboratoire de Microbiologie et Génétique Moléculaire, Université Paul Sabatier-Centre National de la Recherche Scientifique, 118 Route de Narbonne, 31062 Toulouse Cedex, France.
authors.forename: Nelly
authors.initials: N
authors.lastname: Dubarry
authors.forename: Wenli
authors.initials: W
authors.lastname: Du
authors.forename: David
authors.initials: D
authors.lastname: Lane
authors.forename: Franck
authors.initials: F
authors.lastname: Pasta
mesh_terms: D001483:Base Sequence; D042602:Burkholderia cepacia complex / Q000187:drug effects* / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity; D003470:Culture Media; D003550:Cystic Fibrosis / Q000382:microbiology*; D019175:DNA Methylation; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics; D018274:Electroporation / Q000379:methods*; D005798:Genes, Bacterial; D005998:Glycine; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D013096:Spermine / Q000494:pharmacology; D014170:Transformation, Genetic
chemical_list: D003470:Culture Media; D017931:DNA Primers; D004269:DNA, Bacterial; D013096:Spermine; D005998:Glycine