TABLE 1

Strains and plasmids used in this study

Strain or plasmidRelevant characteristic(s)Source or reference
Strains
    S. aureus
        RN4220NCTC8325-4 derivative, antibiotic susceptible, restriction deficient; sau1 hsdR (rK mK+)49
        RN4220/pVPF5RN4220 containing plasmid pVPF5 from S. xylosus VF543
    E. coli
        DH5αK-12 strain; recA1 endA1 hsdR17 (rK mK+)Life Technologies
        AG100K-12 strain50
        AG100AAG100 ΔacrAB::aphA1 (Kmr) mutant; increased susceptibility to antibiotics44
    B. subtilis
        DSM4514Derivative of strain 168 containing plasmid pUB110DSMZ
Plasmids
    pUB110S. aureus plasmid, replicates in B. subtilis; aadD (Kmr Neor)51
    pVPF5S. xylosus pT181 family plasmid; tet(K)43
    pBUS1S. aureus-E. coli shuttle vector; pAMα1 minimum replicon, ColE1, MCS pBluescript II SK (Stratagene), (rrnB T1)5 tet(L)24
    pBUS1-PcapS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, ColE1, promoter of S. aureus type 1 capsule gene 1A (Pcap) upstream of a new MCS with joinable rgs-his6 codons (Pcap-MCS-rgs-his6), (rrnB T1)5 tet(L)This study
    pBUS1-HCS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, sso oriL ColE1, MCS pBluescript II SK (Stratagene), (rrnB T1)5 tet(L)This study
    pBUS1-Pcap-HCS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, sso oriL ColE1 Pcap-MCS-rgs-his6 (rrnB T1)5 tet(L)This study
    pTSSCS. aureus-E. coli shuttle vector; pT181 replicon, ColE1, MCS pBluescript II SK (Stratagene), (rrnB T1)5 tet(L)This study
    pTSSC-PcapS. aureus-E. coli shuttle vector; pT181 replicon, ColE1 Pcap-MCS-rgs-his6 (rrnB T1)5 tet(L)This study
    pTSSCmpTSSC with two silent mutations in the repC sequence to remove HindIII and XbaI sitesThis study
    pTSSCm-PcappTSSC-Pcap with two silent mutations in the repC sequence to remove HindIII and XbaI sitesThis study
    pBJW13S. aureus-E. coli shuttle vector; pAMα1 minimum replicon; ColE1 Pcap erm(44) (MLSBr) (rrnB T1)5 tet(L)26
    pBJW13-HCS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, sso oriL ColE1 Pcap erm(44) (rrnB T1)5 tet(L)This study
    pTJW13S. aureus-E. coli shuttle vector; pT181 replicon, ColE1 Pcap erm(44) (rrnB T1)5 tet(L)This study
    pBJW13-RGS-HisS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, ColE1 Pcap erm(44)-rgs-his6 (rrnB T1)5 tet(L)This study
    pBJW13-HC-RGS-HisS. aureus-E. coli shuttle vector; pAMα1 minimum replicon, sso oriL ColE1 Pcap erm(44)-rgs-his6 (rrnB T1)5 tet(L)This study
    pTJW13-RGS-HisS. aureus-E. coli shuttle vector; pT181 replicon, ColE1 Pcap erm(44)-rgs-his6 (rrnB T1)5 tet(L)This study