TABLE 1.
Strains and plasmids
| Strain or plasmid | Relevant genotype or descriptiona | Source or reference |
|---|---|---|
| Strains | ||
| S. epidermidis | ||
| NIH051475 | 37 | |
| ALC9220 | ΔgraS | This study |
| ALC9221 | ΔgraS WTR | This study |
| ALC9222 | ΔgraS/pEPSA5::vraG; VraG is ectopically overexpressed in ΔgraS background by inducible plasmid pEPSA5 | This study |
| ALC9223 | ΔgraS/pEPSA5::vraFG; VraFG is ectopically overexpressed in ΔgraS background by inducible plasmid pEPSA5 | This study |
| ALC9224 | graS-IMMW2; replacement of strain NIH051475 GraS intramembrane regions with those of S. aureus MW2 over residues 15–34 and 44–64 | This study |
| ALC9225 | ΔEL::Ala9 graS | This study |
| ALC9226 | ΔEL::AYEISVESV graS | This study |
| ALC9227 | ΔEL::DYDFPIDSL graS; replacement of strain NIH051475 9-residue EL with that of S. aureus MW2 | This study |
| ALC9228 | ΔvraG | This study |
| ALC9229 | ΔvraG WT revertant | This study |
| ALC9230 | ΔEL vraG | This study |
| ALC9231 | ΔEL vraG WT revertant | This study |
| ALC9232 | ΔGL vraG | This study |
| ALC9233 | ΔGL vraG WT revertant | This study |
| ALC9234 | ΔGL::Gly10 vraG | This study |
| ALC9235 | vraF-G40A; alanine point mutation of glycine 40 in Walker A motif of VraF | This study |
| ALC9236 | vraF-G40A WT revertant | This study |
| ALC9237 | Strain NIH051475 with C-terminal HA-tagged GraS | This study |
| ALC9238 | Strain NIH051475 with C-terminal HA-tagged EL/DYDFPIDSL GraS | This study |
| ALC9239 | Strain NIH051475 with C-terminal HA-tagged EL/alanine GraS | This study |
| ALC9240 | Strain NIH051475 with N-terminal HA-tagged VraG | This study |
| ALC9241 | Strain NIH051475 with N-terminal HA-tagged ΔEL VraG | This study |
| ALC9242 | Strain NIH051475 with N-terminal HA-tagged ΔGL VraG | This study |
| ALC9243 | Strain NIH051475 with N-terminal HA-tagged GL-Gly10 VraG | This study |
| E. coli | ||
| DC10B | mcrA Δ(mrr-hsdRMS-mcrBC) ϕ80lacZΔM15 ΔlacX74 recA1 araD139 Δ(ara-leu)7697 galU galK rpsL endA1 nupG Δdcm | 31 |
| DTH1 | F− cya-854 ilv-691::Tn10 recA1 endA1 gyrA96 (Nalr) thi-1 hsdR17 spoT1 rfbD1 glnV44(AS) | 38 |
| C41(DE3) | F− ompT gal dcm hsdSB(rB− mB−)(DE3) | Lucigen |
| SKC-N05 | DC10BΩPN25-hsdMS of strain NIH051475 integrated between atpI and gidB genes | This study |
| Plasmids | ||
| pSK236 | Shuttle plasmid; ColE1 E. coli origin with Ampr selection and pC194 staphylococcal origin with Cmr selection | 39 |
| pEPSA5 | Xylose inducible vector; p15A E. coli origin with Ampr selection and pC194 staphylococcal origin with Cmr selection | 34 |
| pMAD | Allelic exchange vector; pBR322 E. coli origin with Ampr selection and pE194ts staphylococcal origin with Ermr selection | 33 |
| pKD46 | λ-red recombineering vector for targeted genetic alterations in E. coli; repA101ts origin, Ampr | 40 |
| pKD4 | Template plasmid for FRT-flanked kanamycin cassette; Ampr and Kanr | 40 |
| pKD46′ | pKD46 harboring E. coli K-12 recA coding sequence under native promoter | This study |
| pKT25 | BacTH vector for N-terminal T25 fusion; p15A origin, Kanr | Euromedex |
| pUT18 | BacTH vector for C-terminal T18 fusion; ColE1 origin, Ampr | Euromedex |
| pKT25-zip | Contains T25 fused leucine zipper of GCN4 | Euromedex |
| pUT18C-zip | Contains T18 fused leucine zipper of GCN4 | Euromedex |
a
Ampr, ampicillin resistance; Cmr, chloramphenicol resistance; Kanr, kanamycin resistance.