TABLE 1.
Bacterial strains, plasmids and oligonucleotide used in this study
| Strain or plasmid | Characteristics | Reference or source |
|---|---|---|
| Strains | ||
| P. aeruginosa | ||
| PDO300 | mucA22 isogenic mutant derived from PAO1 | 18 |
| PDO300ΔalgE | Isogenic algE deletion mutant derived from PDO300 | This study |
| E. coli | ||
| TOP10 | E. coli cloning strain | Invitrogen |
| S17-1 | thi-1 proA hsdR17 (rK− mK+) recA1; tra gene of plasmid RP4 integrated in chromosome | 39 |
| Plasmids | ||
| pGEM-T Easy | Apr, Plac | Invitrogen |
| pBBR1MCS-5 | Gmr; broad-host-range vector; Plac | 23 |
| pBBR1MCS-5:algE | HindIII-BamHI fragment comprising algE inserted into vector pBBR1MCS-5 | This study |
| pBBR1MCS-5:algEDelL7 | algE fragment with 57-bp deletion in region encoding putative surface exposed loop 7 inserted into pBBR1MCS-5 | This study |
| pBBRMCS-5:algEL1FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 189th bp of the ORF | This study |
| pBBRMCS-5:algEL2FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 336th bp of the ORF | This study |
| pBBRMCS-5:algEL3FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 468th bp of the ORF | This study |
| pBBRMCS-5:algEL4FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 582nd bp of the ORF | This study |
| pBBRMCS-5:algEL5FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 711th bp of the ORF | This study |
| pBBRMCS-5:algEL6FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 882nd bp of the ORF | This study |
| pBBRMCS-5:algEL7FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 1035th bp of the ORF | This study |
| pBBRMCS-5:algEL8FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 1269th bp of the ORF | This study |
| pBBRMCS-5:algEL9FLAG | algE fragment with a 24-bp insertion (encoding the FLAG epitope) inserted after the 1428th bp of the ORF | This study |
| pEX100T | Apr Cbr; gene replacement vector containing sacB gene for counterselection | 17 |
| pEX100T:ΔalgEGm | Apr Cbr Gmr; vector pEX100T with SmaI-inserted algE deletion construct | This study |
| pPS865 | Apr Gmr; source of 1,100-bp BamHI fragment comprising aacC1 gene flanked by Flp recombinase target site signal sequences | 17 |
| pPFLP2 | Apr Cbr; broad-host-range vector encoding Flp recombinase | 17 |