TABLE 1.
Strains and plasmids
| Strain or plasmid | Description | Source or reference |
|---|---|---|
| Strains | ||
| B. mallei | ||
| ATCC 23344 | Wild-type strain; polymyxin B resistant, zeocin and kanamycin sensitive | 27 |
| batA KO | Isogenic batA mutant strain of ATCC 23344; resistant to polymyxin B and zeocin, sensitive to kanamycin | This study |
| ilv KO | Isogenic ilvB mutant strain of ATCC 23344; resistant to polymyxin B and kanamycin, sensitive to zeocin | This study |
| B. pseudomallei | ||
| 1026b | Wild-type strain | 148 |
| K96243 | Wild-type strain | 26 |
| B. thailandensis | ||
| DW503 | Laboratory strain; kanamycin sensitive | 143 |
| E. coli | ||
| EPI300 | Strain used for recombinant-DNA manipulations | Epicenter/Illumina |
| S17 | Strain used for conjugational transfer of plasmids to B. mallei; sensitive to polymyxin B, zeocin, and kanamycin | 149 |
| TUNER | Protein expression strain used to purify His- and GST-tagged BatA proteins | EMD Millipore |
| Plasmids | ||
| pBHR1 | Cloning vector; confers resistance to chloramphenicol and kanamycin | MoBiTec |
| pBHR1Δ Dra | pBHR1 containing a 339-nt deletion in the chloramphenicol resistance marker; confers resistance only to kanamycin | 77 |
| pBatA | pBHR1 in which the batA gene of B. pseudomallei K96243 was inserted; confers resistance only to kanamycin | This study |
| pCC1 | Cloning vector; confers resistance to chloramphenicol | Epicenter/Illumina |
| pCCbatA | pCC1 in which the batA gene of B. pseudomallei 1026b was inserted; confers resistance to chloramphenicol | This study |
| pEM7/ZEO | Source of the zeocin resistance cassette; confers resistance to zeocin | Thermo Fisher Scientific |
| pCCbatA.zeo | pCCbatA in which a 1.3-kb portion of the batA ORF was deleted and replaced with a 0.4-kb zeocin resistance cassette; confers resistance to chloramphenicol and zeocin | This study |
| pKAS46 | Gene replacement vector; confers resistance to kanamycin | 150 |
| pKASbatA.zeo | pKAS46 containing the insert from pCCbatA.zeo; confers resistance to kanamycin and zeocin | This study |
| pETcoco-1 | His-tagged protein expression vector; confers resistance to chloramphenicol | EMD Millipore |
| pHisBatA | pETcoco-1 in which a gene fragment encoding amino acids 30–307 of B. mallei ATCC 23344 BatA was inserted; confers resistance to chloramphenicol | This study |
| pGEX4T-2 | GST-tagged protein expression vector; confers resistance to ampicillin | GE Healthcare Life Sciences |
| pGSTBatA | pGEX4T-2 in which a gene fragment encoding amino acids 30–307 of B. mallei ATCC 23344 BatA was inserted; confers resistance to ampicillin | This study |
| pCCilvUP | pCC1 in which a 1-kb PCR product corresponding to the genomic sequence upstream of the B. mallei ATCC 23344 ilvB gene was inserted; confers resistance to chloramphenicol | This study |
| pCCilvDOWN | pCC1 in which a 1-kb PCR product corresponding to the genomic sequence downstream of the B. mallei ATCC 23344 ilvB gene was inserted; confers resistance to chloramphenicol | This study |
| pCCilvΔ | pCC1 containing the genomic sequences located upstream and downstream of the B. mallei ATCC 23344 ilvB gene joined by a unique NheI site; confers resistance to chloramphenicol | This study |
| pUC4K | Source of the kanamycin resistance cassette; confers resistance to kanamycin | GE Healthcare Life Sciences |
| pCCilv.kan | pCCilvΔ in which the kanamycin resistance cassette was inserted into the NheI site | This study |
| pKAS46-ZEO | pKAS46 in which the kanamycin resistance marker was replaced with a zeocin resistance cassette from pEM7/ZEO; confers resistance to zeocin | This study |
| pKASilv.kan | pKAS46-ZEO containing the insert from pCCilv.kan; confers resistance to zeocin and kanamycin | This study |
| pBR322 | Cloning vector; confers resistance to ampicillin | New England Biolabs Inc |
| pJTmcaP | pBR322 in which the gene encoding the M. catarrhalis mcaP gene was inserted; confers resistance to ampicillin | 76 |