title: Stable low-copy-number Staphylococcus aureus shuttle vectors.
abstract: A series of Staphylococcus aureus-Escherichia coli shuttle vectors were constructed which contained the replication and maintenance functions of the S. aureus theta-mode multiresistance plasmid pSK1. The utility of the newly constructed vectors was demonstrated by the successful cloning and expression of several genes that had previously proven difficult to express in S. aureus. Additional vectors which permit the generation of transcriptional and translational fusions to an S. aureus blaZ reporter gene were also produced and subsequently employed to determine the relative strengths in S. aureus of a number of promoters. By utilizing the theta-mode replication functions of pSK1, the shuttle vectors described largely avoided the segregational and structural stability problems frequently encountered with Gram-positive rolling-circle-based vectors. In addition, these plasmids represent vectors which are suitable for the analysis of genes in S. aureus at low copy number.
authors.affiliation: School of Biological Sciences, Macleay Building A12, University of Sydney, New South Wales 2006, Australia.
authors.forename: Steve
authors.initials: S
authors.lastname: Grkovic
authors.affiliation: School of Biological Sciences, Macleay Building A12, University of Sydney, New South Wales 2006, Australia.
authors.forename: Melissa H
authors.initials: MH
authors.lastname: Brown
authors.affiliation: School of Biological Sciences, Macleay Building A12, University of Sydney, New South Wales 2006, Australia.
authors.forename: Kate M
authors.initials: KM
authors.lastname: Hardie
authors.affiliation: School of Biological Sciences, Macleay Building A12, University of Sydney, New South Wales 2006, Australia.
authors.forename: Neville
authors.initials: N
authors.lastname: Firth
authors.affiliation: School of Biological Sciences, Macleay Building A12, University of Sydney, New South Wales 2006, Australia.
authors.forename: Ronald A
authors.initials: RA
authors.lastname: Skurray
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018628:Gene Dosage; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D013211:Staphylococcus aureus / Q000235:genetics*; D014158:Transcription, Genetic; D001618:beta-Lactamases
chemical_list: D001426:Bacterial Proteins; D001618:beta-Lactamases