TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Genotype, relevant phenotype, and/or selection markera | Source or reference |
|---|---|---|
| Strains | ||
| E. coli TOP10 | F−mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 recA1 deoR araD139 Δ(ara leu)7697 galU galK rpsL (Strr) endIA1 nupG | Invitrogen |
| E. coli B178 | W3110 galE sup+ | 12 |
| E. coli MLK217 | B178 lpxL::mini Tn10, Tetr | 12 |
| E. coli MKM10 | MLK217, pMKM.11, Apr, Tetr | This study |
| E. coli MKM11 | MLK217, pMKM.21, Apr, Tetr | This study |
| E. coli MLK1067 | B178 lpxM::Ωcam | 14 |
| E. coli MKM13 | MLK1067, pMKM.11, Apr, Cmr | This study |
| F. tularensis subsp. holarctica strain 1547-57 | Wild-type, human isolate | 36 |
| F. tularensis MKM87 | Strain 1547-57, lpxL::aphA-3, Kmr | This study |
| Plasmids | ||
| pBluescript II SK(−) | Apr, cloning vector | Stratagene |
| pCR2.1TOPO | Apr, Kmr, cloning vector | Invitrogen |
| pCRspec2 | pCR2.1TOPO, aad(9), Apr, Kmr, Spr | This study |
| pFNLTP10grosacB | pFNLTP9 derivative, LVS groEL promoter, sacB, temperature-sensitive origin of Francisella replication, Kmr, Apr | Gift from T. Zhart |
| pMKM.1 | F. tularensis FTT0232c in pCR2.1, Apr, Kmr | This study |
| pMKM.11 | pBluescript II SK(−), EcoRI fragment from pMKM.1 containing FTT0232c, Kmr | This study |
| pMKM.16 | pCR2.1TOPO, FTT0232c and flanking sequence without consensus sequence regions, Apr, Kmr | This study |
| pMKM.17 | pMKM.16, aphA-3, Apr, Kmr | This study |
| pMKM.2 | pCR2.1TOPO, FTT0231c, Apr, Kmr | This study |
| pMKM.21 | pBluescript II SK(−), EcoRI fragment from pMKM.1 containing FTT0231c, Kmr | This study |
| pMKM.219c | 7.5-kb plasmid containing F. tularensis temperature-sensitive origin of replication, LVS groEL promoter upstream of sacB, E. coli origin of replication, aad(9), Spr | This study |
| pMKM.223 | pMKM.219c, FTT0232c::aphA-3, Spr, Kmr | This study |
| pSPECR | pCRBlunt (Invitrogen), aad(9), Spr | 57 |
| pUC18K | aphA-3, Kmr | 33 |
a
Spr, Apr, and Kmr, resistance to spectinomycin, ampicillin, and kanamycin, respectively.