TABLE 1.
Bacterial strains and plasmids used
| Strain or plasmid | Relevant characteristic and/or genotypea | Comments (reference or source) |
|---|---|---|
| Strains | ||
| S. aureus | ||
| RN4220 | Restriction-deficient mutagenized RN450 | Shuttle plasmid host (16) |
| 450M | Cms Ems Gms Mcr (HE) Tcs Bla− | RN450 transformed with COL mec region DNA (1) |
| N315P | Cms Emr Gms Mcr (HE) Tcs Bla− | Intact mecR1-mecI; clinical isolate from Japan (13) |
| 67-0 | Mcr Emr (HE) Bla− | Clinical isolate from California, 1984 (5) |
| E. coli | ||
| TB1 | recA+lacIqlacZΔM15 | Host for pUC vectors (26) |
| DH5α | F−recA1 hsdR17 φ80dlacZM15Δ(lacZYA-argF) endA1 deoR gyr A96 thi-1 relA supE44 | Recombination-deficient host for pUC derivatives (Gibco) |
| Plasmids | ||
| pUC19 | Apr; 2.7 kb | General-purpose cloning vector (26) |
| pGEX-2T | Apr; 4.8 kb | Vector for the expression of GST-protein fusions (Pharmacia) |
| pSK950 | Spr Tcr Emr ts; 10.4 kb | E. coli-S. aureus shuttle vector for integration of sequences into the S. aureus lipase gene with φL54a att site. The original pCL84 plasmid was modified by addition of pE194 at the PstI site (17). |
| pG0630 or pG0631 | Spr Tcr Emr ts; 16.4 kb | A 2.4-kb fragment from N315P(pG0630) or 67-0(pG063), containing the mecR1-mecI intergenic region with divergent mecR1-mecI and mecA promoters, and the first 50 bases of the mecA coding sequence cloned at EcoRI-BamHI sites of pSK950. A 3.3-kb fragment containing promoterless lacZ was then cloned at the BamHI site to generate a mecA-lacZ transcriptional fusion (this study). |
a
Abbreviations: Ap, ampicillin; Bla, β-lactamase; Cm, chloramphenicol; Em, erythromycin; Gm, gentamicin; HE, heterotypic; Mc, methicillin; MCS, multiple cloning site; Sp, spectinomycin; Tc, tetracycline; ts, temperature sensitive.