TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Relevant genotype or propertiesa | Reference or source |
|---|---|---|
| E. coli strains | ||
| DH5α | F− φ80lacZΔM15 Δ(lacZYA-argF)U169 deoR recA1 endA1 hsdR17(rK− mK+) phoA supE44 | 14 |
| SM10(λpir) | Kmr; thi-1 thr-1 leuB26 tonA21 lacY1 supE44 recA integrated RP4-2 Tcr::Mu aphA+ (RP4-2 is RP4 ΔTn1) | 17 |
| RHO1 | Kms; SM10(λpir) ΔaphA::FRT | This study |
| RHO2 | Kmr; SM10(λpir) Δasd::FRT | This study |
| RHO3 | Kms; SM10(λpir) Δasd::FRT ΔaphA::FRT | This study |
| B. pseudomallei strains | ||
| 1026b | Wild-type strain, clinical isolate | 7 |
| K96243 | Sequenced prototype strain, clinical isolate | 10 |
| Plasmids | ||
| pBAD-I-sceI | Apr; source of araC-PBAD-I-sceI expression cassette | 33 |
| pBADSceb | Zeor; I-SceI expression vector with pRO1600(Ts) replicon | This study |
| pEX100T | Apr; source of I-SceI sites, lacZα, and pMB9 ori | 25 |
| pEXKm4b | Kmr; pEX100T with pFKM2 nptII gene, E. coli gusA reporter gene, and a synthetic multiple cloning site | This study |
| pEXKm5b | Kmr; pEXKm4 with B. pseudomallei optimized sacB gene region from pMo130 | This study |
| pFKM2 | Apr Kmr; source of nptII gene | 4 |
| pMo130 | Kmr; source of sacB gene optimized for B. pseudomallei | 9 |
| pPS2382 | Apr Kmr; pCR2.1 TA cloning vector (Invitrogen) with B. pseudomallei fabD gene | This study |
| pPS2383 | Apr Kmr; pPS2382 with fabD(Ts) allele | This study |
| pPS2592 | Kmr; pEXKm5 with fabD(Ts) allele fragment from pPS2383 | This study |
a
Abbreviations: Ap, ampicillin; Km, kanamycin; r, resistant; s, susceptible; Tc, tetracycline; Tp, trimethoprim; Zeo, zeocin.
b
Details on the construction of pBADSce, pEXKm4, pEXKm5, and intermediary plasmids are provided in the supplemental material.