TABLE 1.
Strains and plasmids used in this study
| Strains/plasmids | Description | Source or reference |
|---|---|---|
| Strains | ||
| P. aeruginosa | ||
| 1244 | Wild type, IATS O7 | Ref. 39 |
| 1244N3 | rpoN (TcR) | Ref. 27 |
| 1244.47 | pilAO (HgR) | This study |
| 1244.2024 | pilAOBCD (KmR) | This study |
| PA103 | Wild type, IATS O11 | Ref. 40 |
| E. coli | ||
| XL1-Blue | recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac | Stratagene |
| XL10-Gold | endA1 supE44 thi-1 recA1 gyrA96 relA1 lacHte | Stratagene |
| DH5α | supE44 hsdR17 recA1 endA1 gyrA96 thi-1 relA1 | Invitrogen |
| HB101 | supE44 hsdS20 recA13 ara-14 proA2 lacY1 galK2 rpsL20 xyl-5 mtl-1 | Ref. 41 |
|
| ||
| Plasmids | ||
| pUCP26 | 4.9 kb pUC18-derived broad host-range cloning vector, TcR | Ref. 42 |
| pUCP26pilO | pUCP26 with 1244 pilO | This study |
| pMMB66EH | 8.8-kb broad host range expression vector, ApR/CbR | Ref. 43 |
| pPAC46 | pMMB66EH with 1244 pilAO | Ref. 2 |
| pPAC202∷Tn501-13 | A cosmid subclone containing a transposon insertion in 1244 pilA | Ref. 21 |
| p1244 S148T | pPAC46 with coding sequence for Ser148 of pilA mutated to Thr | This study |
| p1244 A149 | pPAC46 with the addition of an Ala and a stop codon after Ser148 of pilA | This study |
| pSD5 | pMMB66EH with PA103 pilA | This study |
| p103 P144S | pSD5 with the Pro144 codon of pilA mutated to Ser | This study |
| p103 PKS | pSD5 with codons for Asn142, Glu143, and Pro144 of pilA mutated to Pro, Lys, and Ser, respectively | This study |
| pHYBRID | pMMB66EH with the chimeric pilA coding for the first 123 residues of PA103 pilin, a Gly at residue 124, and the last 31 residues are those of 1244 pilin; this plasmid also contains pilO | This study |
| pHYB-O | pMMB66EH with the chimeric pilA from pHYBRID; this plasmid does not contain pilO | This study |
| p1244 C127A | pPAC46 with coding sequence for Cys127 of pilA mutated to Ala | This study |
| p1244 C145A | pPAC46 with coding sequence for Cys145 of pilA mutated to Ala | This study |
| p1244Δ135–139 | pPAC46 with coding sequence for residues 135–139 of pilA (AWKPN) deleted | This study |
| p1244Δ135–144 | pPAC46 with coding sequence for residues 135–144 of pilA (AWKPNYAPAN) deleted | This study |
| p1244Δ128–144 | pPAC46 with coding sequence for residues 128–144 of 1244 pilA (KITKTPTAWKPNYAPAN) deleted | This study |
| p103 INS136 | pSD5 with coding sequence for residues 135–139 of 1244 pilA (AWKPN) inserted at amino acid positions 136–141 of PA103 pilA with Pro144 (now Pro149) mutated to Ser | This study |
| p1244 S147 | pPAC46 with mutant pilA that had coding sequence for Lys 147 mutated to Ser and Ser 148 mutated to a stop codon | This study |
| p1244 S149 | pPAC46 with 1244 pilA that had the Ser148 codon mutated to Ala and the addition of Ser 149 followed by a stop codon | This study |
| p1244 S150 | pPAC46 with mutant pilA that had coding sequence for Ser148 mutated to Ala, an Ala at position 149, a Ser at position 150 followed by a stop codon | This study |
| p103 S146 | pSD5 with coding sequence for two Ala inserted into PA103 pilA at positions 144 and 145 terminating in Ser146 | This study |