Table 1.
Strains, plasmids, and primers used in this study
| Strain, plasmid, or primer | Description | Note |
|---|---|---|
| Strains | ||
| P. aeruginosa PAO1 | WT | ATCC |
| E. coli BL21(DE3) | F− ompT hsdSB (rB− mB−) gal dcm (DE3) | Invitrogen |
| E. coli SM10 λpir | thi-1 thr leu tonA lacY supE recA::RP4-2-Tc::Mu Km λpir | Yonsei Univ. |
| PW7731 | thiE transposon mutant, lacZbpO2q3B12 | (53) |
| ΔthiL | PAO1 with thiL(PA4051) in-frame gene deletion | This study |
| Plasmidsa | ||
| pET28b(+) | E. coli expression vector, KmR | Novagen |
| pCVD442-Gm | sacB suicide vector from pUM24, GmR | Yonsei Univ. |
| pBSP II SK(−) | Broad-host-range expression vector, AmpR | (54) |
| pETthiL | PAO1 thiL gene inserted in pET28b(+) | This study |
| pthiL | PAO1 thiL gene inserted in pBSP II SK(−) | This study |
| Primersb | ||
| thiL (PA4051) | Forward, TAGGTTGGATCCCTGCGGCTGTCCACCTACGAGC | BamHI |
| Forward, TAGGTTCATATGGGTGAGTTCGAGCTGATCCGCC | NdeI | |
| Reverse, TAGGTTAAGCTTTCAGTCACGTTGGGTTCCGAAATGTTGG | HindIII | |
| nusB (PA4052) | Forward, GTTATAGCATGCGTGAGCAATCAAGACAGCGGC | SphI |
| Reverse, ATTAATCGAGCTCTCAGCGCTTGCCGCCG | SacI | |
| pgpA (PA4050) | Forward, GTTGTTGAGCTCGTGACTGAGCATCCTGATCAG | SacI |
| Reverse, GTTGTTCCCGGGTCAGACCAGCCAGTG | SmaI |
aKmR, kanamycin resistance; GmR, gentamycin resistance; AmpR, ampicillin resistance.
bRestriction sites are underlined.