Table 1.

Strains, plasmids, and primers used to make the deletion mutants

Strain, plasmid, or primerGenotype, relevant phenotype,a or sequenceSource or reference
A. baumannii strains
    ATCC 17978WTATCC
    R2, R3, R5, R6, R7, and R9Polymyxin-resistant mutants spontaneously derived on gradient plates from ATCC 17978This study (Table 3)
E. coli strains
    TOPO10F mcrA D(mrr-hsdRMS-mcrBC) φ80lacZ M15 D lacX74 recA1 araD 139 D (ara-leu)7697 galU galK rpsL (strR) endA1 nupGInvitrogen
    DH5αInvitrogen
    W3110Wild type; F λE. coli Genetic Stock Center (Yale)
    WD101W3110 constitutive pmrA mutant (polymyxin resistant)13
Plasmids
    pCR-Blunt II-TOPOPCR cloning vector; KmrInvitrogen
    pUCK4Source of Kmr kanamycin cassette genePharmacia
    pEX18 AmpSuicide plasmid; Ampr; sacB14
    pWH1266E. coli-Acinetobacter shuttle plasmid; Ampr CBr Tetr15
    pABNpWH1266 harboring pmrAB from the WT; CBrThis study
    pAB25pWH1266 harboring pmrAB from R2; CBrThis study
    pAB10pWH1266 harboring pmrAB from R5; CBrThis study
Primers
    AB pmrA NFCAGCAGTTTCTGTGCCATGTThis study
    KanPmrA NRTGAGACACAACGTGGCTTTCCCATCCATCATAGGCAATCCThis study
    KanPmrA CFATTACGCTGACTTGACGGGAACCGTTTGGGGCAATCATAThis study
    AB pmrA CRTTTTGCATAGCCAAGTTGACCThis study
    nest-A-FCGGGTATGCCACGTGTAGATThis study
    nest-A-RAATGCAGTCACAGGTGTTCGThis study
    AB pmrB NFGGTCATCAACCTCAACTGGThis study
    KanPmrB NRTGAGACACAACGTGGCTTTCGGGTGCTCAGCTGTTCTTTCThis study
    KanPmrB CFATTACGCTGACTTGACGGGATTTACATGAAACAAGAGCGTGAThis study
    AB pmrB CRTTAAAGCGACCATTGGCATTThis study
    nestB-FGGTGGAATGGGTCAATAACGThis study
    nestB-RGAGTTCCGTGGTGCAGACTTThis study
    C0FTGATAAATTACGTGCGACTGCThis study
    KC1R1TGAGACACAACGTGGCTTTCTTCAGCATCGTGAGTGACTACAThis study
    KC2F2ATTACGCTGACTTGACGGGAGTGCCCATGTAAACTAAAGCGThis study
    YnewRTTGCCAATGCTAAAAGCTGAThis study
    C1FGCGTATTCGCCATGCTTATTThis study
    C2RGCTTCACGAACCCTCTCTTGThis study
  • a Antibiotic resistance phenotypes: Kmr, kanamycin resistant, Ampr, ampicillin resistant for E. coli; CBr, carbenicillin resistant for A. baumannii; Tetr, tetracycline resistant.