TABLE 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Descriptiona | Source or reference(s) |
|---|---|---|
| E. coli | ||
| DH5α | F− φ80 lacZΔM15 Δ(lacZYA-argF) recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 deoRU169 | Invitrogen |
| HB101 | F− Δ(gpt-proA)62 leuB6 glnV44 ara-14 galK2 lacY1 Δ(mcrC-mrr) rpsL20 (Strr) xyl-5 mtl-1 recA13 | 57 |
| TOP10 | F−mcrA Δ (mrr-hsdRMS-mcrBC) φ80 lacZΔM15 Δ lacX74 deoR recA1 araD139 Δ (ara-leu)7697 galU galK rpsL (Strr) endA1 nupG | Invitrogen |
| SM10 | Mobilizing strain, RP4 genes integrated in chromosome, Kmr | 62 |
| B. cenocepacia | ||
| Pc715j | CF respiratory isolate; Orn, Pch, SA | 19, 40 |
| Pc715jorbA::tp | orbA::Tpr derivative of Pc715j; Tpr | This study |
| Pc715jfptA::tp | fptA::Tpr derivative of Pc715j; Tpr | This study |
| K56-2 | CF respiratory isolate; Orn, SA | 19, 39 |
| K56orbA::tp | orbA::Tpr derivative of K56-2; Tpr | 63 |
| Plasmids | ||
| pRK2013 | ColE1 tra(RK2)+; Kmr | 23 |
| pCC524T | pEX18Tc with a 3-kb EcoRI-BglII orbA fragment from K56-2 containing a 0.6-kb Tpr cassette inserted into the orbA gene; Tcr Tpr | 63 |
| pCRR2.1TOPO | Cloning vector for PCR products; Kmr Apr | Invitrogen |
| pEX18Tc | Suicide vector; sacB Tcr | 31 |
| p34E-Tp | Source of trimethoprim resistance cassette; Tpr | 20 |
| pEKH103 | pCRR2.1TOPO containing a 2.35-kb fptA PCR product; Apr Kmr | This study |
| pEKH107 | pEKH103 with a 0.67-kb SalI fragment containing the Tpr cassette from p34E-Tp inserted into the SalI site in the fptA gene; Apr Kmr Tpr | This study |
| pEKH113 | pEX18Tc with a 2.2-kb HindIII-PstI fptA::tp fragment from pEKH107; Tcr Tpr | This study |
a
Tc, tetracycline; Tp, trimethoprim; Km, kanamycin; Ap, ampicillin; Orn, ornibactin; Pch, pyochelin; Str, streptomycin; CF, cystic fibrosis.