title: Eukaryotic expression vectors that replicate to low copy number in bacteria: transient expression of the Menkes protein.
abstract: A set of low copy number plasmid vectors for mammalian gene expression has been constructed. These vectors are derived from the previously described bacterial low copy number expression vectors, pWSK29 and pWKS30, which are present at six to eight copies per cell. The new plasmids also have the following useful properties: (1) they contain antibiotic resistance markers for the selection of stable mammalian cell lines; (2) they have either constitutive or inducible promoters; (3) a chimeric intron, for enhancing gene expression, is present; (4) they contain unique cloning sites; (5) they have an SV40 polyadenylation signal, and a subset of the vectors have an SV40 origin of replication for episomal replication and transient gene expression. A cDNA encoding the Menkes disease protein was cloned into two of these vectors, and transient expression studies in COS-7 cells showed that both constitutive and inducible expression was possible. This set of expression vectors will provide a useful tool for the manipulation, in Escherichia coli, of mammalian genes or cDNAs that are unstable in the high copy number vectors that are currently available.
authors.affiliation: The Murdoch Institute for Research into Birth Defects, Royal Children's Hospital, Parkville, Victoria, 3052, Australia.
authors.forename: S L
authors.initials: SL
authors.lastname: Fontaine
authors.forename: S D
authors.initials: SD
authors.lastname: Firth
authors.forename: P J
authors.initials: PJ
authors.lastname: Lockhart
authors.forename: J A
authors.initials: JA
authors.lastname: Paynter
authors.forename: J F
authors.initials: JF
authors.lastname: Mercer
mesh_terms: D000251:Adenosine Triphosphatases / Q000096:biosynthesis* / Q000235:genetics*; D000818:Animals; D001419:Bacteria / Q000235:genetics*; D019556:COS Cells; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics*; D027682:Cation Transport Proteins; D000073840:Copper-Transporting ATPases; D004261:DNA Replication; D005057:Eukaryotic Cells / Q000382:microbiology*; D018628:Gene Dosage; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D011993:Recombinant Fusion Proteins
chemical_list: D002352:Carrier Proteins; D027682:Cation Transport Proteins; D011993:Recombinant Fusion Proteins; D000251:Adenosine Triphosphatases; C081184:ATP7A protein, human; D000073840:Copper-Transporting ATPases