TABLE 1.
Bacterial strains and plasmids used in this study
| Bacterial strain or plasmid | Phenotype, genotype, and/or descriptiona | Source or reference |
|---|---|---|
| P. aeruginosa strains | ||
| PAO1 | Alg−, prototroph | P. Phibbs |
| PAO1 ΔalgB | Alg−, PAO1 in-frame deletion of algB (PA5483) | This study |
| PAO1 ΔalgU | Alg−, PAO1 in-frame deletion of algU (PA0762) | This study |
| PAO1 ΔkinB | Alg+, PAO1 in-frame deletion of kinB (PA5484) | This study |
| PAO1 ΔrpoN | Alg−, PAO1 in-frame deletion of rpoN (PA4462) | This study |
| PAO1 kinB::aacC1 | Alg+, PAO1 kinB::Gmr | This study |
| PAO1 kinB::aacC1 ΔalgU | Alg−, PAO1 kinB::aacC1 in-frame deletion of algU (PA0762) | This study |
| PAO1 ΔalgB ΔkinB | Alg−, PAO1 in-frame deletion of algB (PA5483) and kinB (PA5484) | This study |
| PAO1 kinB::aacC1ΔalgW | Alg−, PAO1 kinB::aacC1 in-frame deletion of algW (PA4446) | This study |
| PAO1 kinB::aacC1 ΔrpoN | Alg−, PAO1 kinB::aacC1 in-frame deletion of rpoN (PA4462) | This study |
| E. coli strains | ||
| DH5α | F− φ80dlacZΔM15 Δ(lacZYA-argF)U169 deoR recA1 endA1 hsdR17(rK− mK+) phoA supE44 λ−thi-1 gyrA96 relA1 | Lab strain |
| TOP10 | DH5α derivative | Invitrogen |
| SM10/λpir | thi recA thr leu tonA lacY supE RP4-2-Tc::Mu1::pir Kmr | Lab strain |
| Plasmids | ||
| pRK2013 | Kmr Tra Mob ColE1 | 14 |
| pFAC | Mini-himar1 mariner transposon in Pseudomonas suicide plasmid; Apr Gmr | 53 |
| pCR4-TOPO | TA cloning vector; 3.9 kb; Apr Kmr | Invitrogen |
| pUS56 | algB45 in pTrcHisA BamHI-EcoRI | 28 |
| pHERD20T | pUCP20T Plac replaced by 1.3-kb AflII-EcoRI fragment of araC-PBAD cassette | 41 |
| pHERD20T-algU | algU (PA0762) from PAO1 in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-algB | algB from PAO1in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-algB45 | algB45 from pUS56 in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-algW | algW (PA4446) from PAO1in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-MPDZalgW | algW with partial PDZ domain in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-ΔPDZalgW | algW with complete deletion of PDZ domain in pHERD20T EcoRI/HindIII | This study |
| pHERD20T-kinB | kinB from PAO1 in pHERD20T KpnI/HindIII | This study |
| pHERD20T-HA-mucA | N-terminally tagged HA-mucA in pHERD20T EcoRI/HindIII | This study |
| pUCP20T-PBAD-rpoN | araC-PBAD-rpoN fusion in pUCP20 XbaI/HindIII | This study |
| MiniCTX-lacZ | Gene delivery vector for inserting genes at the CTX phage att site on the P. aeruginosa chromosome; Tcr | 20 |
| MiniCTX-PalgU-lacZ | Complete PalgU promoter (541 bp upstream of ATG) EcoRI/HindIII fused with lacZ for integration at the CTX phage att site in P. aeruginosa | This study |
| MiniCTX-PalgD-lacZ | Complete PalgD promoter (1,525 bp upstream of ATG) HindIII/BamHI fused with lacZ for integration at the CTX phage att site in P. aeruginosa | This study |
| pEX100T | Pseudomonas suicide vector, sacB oriT Cbr | 47 |
| pEX100T-NotI | Pseudomonas suicide vector with NotI restriction site fused into SmaI of pEX100T, sacB oriT Cbr | This study |
| pEX100T-ΔalgU | A 2.5-kb fragment flanking algU gene fused with pEX100T-NotI with in-frame deletion of algU with only 24 bp left coding for 8 amino acids of algU | This study |
| pEX100T-ΔalgB | A 1.5-kb fragment flanking algB gene fused with pEX100T-NotI with in-frame deletion of algB | This study |
| pEX100T-ΔalgBΔkinB | A 1.5-kb fragment flanking algB gene fused with pEX100T-NotI with in-frame deletion of algB and kinB | This study |
| pEX100T-ΔalgW | A 1.4-kb fragment flanking algW gene fused with pEX100T-NotI with in-frame deletion of algW | 43 |
| pEX100T-ΔkinB | A 2.5-kb fragment flanking kinB gene fused with pEX100T-NotI with in-frame deletion of kinB | This study |
| pEX100T-ΔrpoN | A 1.9-kb fragment flanking rpoN gene fused with pEX100T-NotI with in-frame deletion of rpoN | This study |
a
Alg−, nonmucoid phenotype; Alg+, mucoid phenotype.