TABLE 1.
Yeast and bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant genotypea | Source of reference |
|---|---|---|
| S. cerevisiae AH109 | MATatrp1-901 leu2-3, 112 ura3-52 his3-200 gal4Δ gal80Δ LYS2::GAL1UAS-GAL1TATA-HIS3 GAL2UAS-GAL2TATA-ADE2 URA3::MELIUAS-MEL1TATA-lacZ | Clontech |
| E. coli DH5α | hsdR recA lacZYA φ80 lacZΔM15 | Gibco-BRL |
| P. aeruginosa | ||
| PAK | Wild-type clinical isolate | D. Bradley |
| PAK-M | PAK flgM::Gmr | This study |
| PAK-N1G | PAK rpoN::Gmr | 18 |
| PAK-Q | PAK fleQ::Gmr | 3 |
| MS540 | PAK fliA::Gmr | 33 |
| Plasmids | ||
| pCR2.1 | Cloning vector, Ampr Kanr, LacZα | Invitrogen |
| pCR2.1-M | pCR2.1 containing a 2.0-kb SstI/HindIII fragment including the flgM gene from PAK | This study |
| pCR2.1-MG | pCR2.1-M with a Gmr gene inserted into a unique BamHI site of the flgM gene | This study |
| pET15bVP | Expression vector, T7 promoter, His tag coding sequence, Ampr, pBR322 origin, contains a broad-host-range origin of replication oriV | 3 |
| pET15bVP-M | pET15bVP with a 324-bp NdeI/BamHI PCR fragment of the PAK flgM gene | This study |
| pET15bVP-A | pET15bVP with a 744-bp NdeI/BamHI PCR fragment of the PAK fliA gene | This study |
| pMMB67HE | Broad-host-range cloning vector, tac promoter, Carbr | 30 |
| pMMB67HE-M | pMMB67HE with a 1.2-kb EcoRI/SstI fragment from pCR2.1-FM containing the flgM gene | This study |
| pDN19lacΩ | Promoterless lacZ oriV oriT Tcr Strr Ω fragment | 34 |
| pDN19lacΩ-M | pDN19lacΩ with a 614-bp EcoRI/BamHI fragment of the flgM gene promoter region | This study |
| pMS565 | pDN19lacΩ containing the fliA promoter region | 33 |
| pPT269 | pDN19lacΩ containing the fliC promoter region | 34 |
| pGADT7 | Cloning vector, GAL4768-881 AD, LEU2, Ampr, HA epitope tag | Clontech |
| pGBKT7 | Cloning vector, GAL41-147 DNA-BD, LEU2, Kanr, c-Myc epitope tag | Clontech |
| pGADT7-M | PAK flgM gene of 324-bp inserted as a PCR product into the NdeI/BamHI sites of pGADT7 | This study |
| pGADT7-A | PAK fliA gene of 744-bp inserted into the NdeI/BamHI sites of pGBKT7 | This study |
| pGBKT7-M | PAK flgM gene of 324-bp inserted as a PCR product into the NdeI/BamHI sites of pGBKT7 | This study |
| pGBKT7-A | PAK fliA gene of 744-bp inserted into the NdeI/BamHI sites of pGBKT7 | This study |
a
Gmr, gentamicin resistance; Kanr, kanamycin resistance; Carbr, carbenicillin resistance; Strr, streptomycin resistance; Tcr, tetracycline resistance; Ampr, ampicillin resistance; HA, hemagglutinin.