title: A novel plasmid (pEMCJH03) isolated from moraxella catarrhalis possibly useful as a cloning and expression vector within this species.
abstract: A preliminary screening study of six Moraxella catarrhalis isolates from primary school children in the Netherlands identified a small 3.5 kb plasmid (pEMCJH03), containing four open reading frames, which encoded three mobilizing and one replicase protein. Insertion of a kanamycin containing transposon (yielding pEMCJH04) allowed selection and isolation of the plasmid in Escherichia coli. Natural transformation of pEMCJH04 into M. catarrhalis was successful for 25% (3/12) of non-isogenic isolates, with no link between (lack of) transformability and genetic lineage or (lack of) transformability and complement phenotype, though the transformation efficiency was found to be rather low at approximately 615CFU/microg (range=60-1040CFU/microg ). This is only the second publication detailing a plasmid isolated from this important respiratory pathogen, and the ability to clone and express foreign proteins in M. catarrhalis using pEMCJH04 could help in the development of a vaccine expression vector, as well as providing a useful tool for studying promoter activity and in complementation studies of gene knockout mutants.
authors.affiliation: Department of Medical Microbiology and Infectious Diseases, Erasmus MC (Erasmus University Medical Center), Dr. Molewaterplein 40, 3015 GD Rotterdam, The Netherlands. j.hays@erasmusmc.nl
authors.forename: John P
authors.initials: JP
authors.lastname: Hays
authors.forename: Kimberly
authors.initials: K
authors.lastname: Eadie
authors.forename: Cees M
authors.initials: CM
authors.lastname: Verduin
authors.forename: Henri
authors.initials: H
authors.lastname: Verbrugh
authors.forename: Alex
authors.initials: A
authors.lastname: van Belkum
mesh_terms: D001426:Bacterial Proteins; D001483:Base Sequence; D002648:Child; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D005822:Genetic Vectors; D006801:Humans; D007612:Kanamycin / Q000737:chemistry; D008969:Molecular Sequence Data; D001936:Moraxella catarrhalis / Q000235:genetics* / Q000472:pathogenicity; D016254:Mutagenesis, Insertional; D009426:Netherlands; D016366:Open Reading Frames; D010957:Plasmids / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D012091:Repetitive Sequences, Nucleic Acid; D012641:Selection, Genetic; D013045:Species Specificity; D014169:Transformation, Bacterial
chemical_list: D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D007612:Kanamycin