TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Relevant genotype or phenotypea | Reference or source |
|---|---|---|
| Strains | ||
| E. coli | ||
| DH10B | F− φ80dlacZΔM15 recA1 | Gibco, Gaithersburg, Md. |
| BL21(DE3) | F−ompT gal [dcm] [lon] hsdSB (rB− mB−), with DE3 | Novagen, Madison, Wis. |
| S. aureus | ||
| RN4220 | NCTC8325-4 r− m+ (restriction minus, modification plus) | 26 |
| BB255 | Essentially the same as NCTC8325 | 5 |
| 8325-4 | NCTC8325, cured of known prophages | 34 |
| RN6390 | Derivative of NCTC8325 that maintains its hemolytic pattern when propagated on sheep erythrocytes | 35 |
| COL | mec, high-Mcr clinical isolate, Tcr | 25 |
| Newman | Clinical isolate, high level of clumping factor (ATCC 25904) | 18 |
| IK181 | BB255 ΔrsbUVWsigB Emr | 28 |
| IK183 | COL ΔrsbUVWsigB Emr | 28 |
| IK184 | Newman ΔrsbUVWsigB Emr | 28 |
| GP268 | BB255 (rsbUVWsigB)+-tetL Tcr | This study |
| GP269 | 8325-4 (rsbUVWsigB)+-tetL Tcr | This study |
| MB25 | RN4220 asp23+ (asp23P::luc+)-pEC-ermB Emr | This study |
| MB32 | Newman asp23+ (asp23P::luc+)-pEC-ermB Emr | This study |
| MB33 | BB255 asp23+ (asp23P::luc+)-pEC-ermB Emr | This study |
| MB49 | BB255 (rsbUVWsigB)+-tetL asp23+ (asp23P::luc+)-pEC-ermB Tcr Emr | This study |
| MB61 | RN4220 asp23+ (asp23P::luc+)-pBT-tetL Tcr | This study |
| MB69 | Newman ΔrsbUVWsigB asp23+ (asp23P::luc+)-pBT-tetL Tcr Emr | This study |
| MB90 | BB255 ΔrsbUVWsigB asp23+ (asp23P::luc+)-pBT-tetL Tcr Emr | This study |
| Plasmids | ||
| pET-24b(+) | Kmr; expression vector | Novagen |
| pSP-luc+ | Apr; firefly luciferase casette vector | Promega |
| pBC SK+ | Cmr; cloning vector | Stratagene |
| pAW8 | Tcr; pAMα1 origin and tetL gene of pHY300PLK, ColE1 origin | A. Wada, unpublished data |
| pBT | Tcr; 1.6-kb PCR fragment of tetL gene of pHY300PLK into Alw26I-digested pBC SK(+) | This study |
| pEC1 | Apr; Emr; 1.45-kb ClaI ermB fragment of Tn551 in pUC18 | 9 |
| pIK6 | Apr; 6.6-kb PstI-EcoRI sigB fragment from strain 8325 in pUC18 | 27 |
| pPG11 | Apr; Tcr; 252-bp MluI-BstXI fragment of rsbU from strain COL replacing the corresponding region of the 6.6-kb PstI-EcoRI sigB fragment from strain 8325 in pUC19 | This study |
| pETasp23 | 510-bp PCR fragment of asp23 from strain 8325 in pET24b | This study |
| pETrsbUCOL | 1-kb PCR fragment of rsbU from strain COL in pET24b | This study |
| pETsigB | 770-bp PCR fragment of sigB from strain 8325 in pET24b | This study |
| pSPasp23P | 1.1-kb PCR fragment of asp23 promoter from strain COL in pSP-luc+ | This study |
| pECasp23P::luc+ | 2.7-kb KpnI-EcoRI asp23P::luc+ fragment of pSPasp23P in pEC1 | This study |
| pBTasp23P::luc+ | 2.7-kb KpnI-EcoRI asp23P::luc+ fragment of pSPasp23P in pBT | This study |
a
Abbreviations are as follows: Apr, ampicillin resistant; Cmr, chloramphenicol resistant; Emr, erythromycin resistant; Kmr, kanamycin resistant; Mcr, methicillin resistant; Tcr, tetracyclin resistant.