TABLE 1.
Strains, plasmids, and oligonucleotides used in this study
| Strain, plasmid, or oligonucleotide | Relevant characteristic(s) or sequence (5′ to 3′) | Source or reference |
|---|---|---|
| Strains | ||
| Escherichia coli | ||
| EC100D+pir | supE44 ΔlacU169 (ϕ80 lacZΔM15) hsdR17 recA1 endA1 gyrA96 thi-1 relA1 (λpirR6K) | Epicenter |
| SM10λpir | thi-1 thr leu tonA lacY supE recA::RP4-2-Tc::Mu kmR (λpirR6K) | Laboratory collection |
| ER2566 | F− glnV44(AS) galK2(Oc) rpsL704(strR) xylA5 mtl-1 argE3(Oc) thiE1 tfr-3 λ DE3 λ sBamHIo ΔEcoRI-B int::(lacI::PlacUV5::T gene1) i21 Δnin5 | New England BioLabs |
| Vibrio cholerae | ||
| JB3 | V. cholerae O1 El Tor strain N16961, Smr | Laboratory collection |
| JB58 | V. cholerae O1 El Tor strain N16961, Smr, ΔlacZ | Laboratory collection |
| XBV222 | JB58 ΔleuO | 11 |
| DT733 | JB58 ΔtoxRS | 11 |
| JB461 | JB3 ΔtoxRS | 30 |
| XBV302 | JB58 ΔalmE | This study |
| XBV532 | V. cholerae O1 El Tor strain N16961, Smr, ΔlacZ ΔompU | 17 |
| XBV534 | XBV532 ΔleuO | This study |
| XBV537 | XBV532 ΔtoxRS | This study |
| Plasmids | ||
| pTL61T | lacZ transcriptional reporter plasmid, Cbr | 31 |
| pXB266 | pTL61T containing the leuO promoter region | 11 |
| pVA289 | pTL61T containing the carRS promoter region | This study |
| pXB340 | pTL61T containing the carRS promoter region | This study |
| pXB342 | pTL61T containing the almEFG promoter region | This study |
| pBAD18Km | Arabinose-regulated expression plasmid, Kmr | 32 |
| pXB269 | pBAD18Km expressing VC2486 and leuO | 11 |
| pBAD33 | Arabinose-regulated expression plasmid, Cmr | 32 |
| pVA126 | pBAD33 expressing leuO | 19 |
| pXB302 | pBAD33 expressing toxRS | 10 |
| pMAL-c2 | IPTG-inducible expression vector for fusion of proteins to MBP and cytoplasmic expression, Cbr | New England BioLabs |
| pVA175 | pMAL-c2 expressing leuO | 19 |
| pWM91 | Suicide plasmid vector used for allelic exchange, Cbr | 33 |
| pWM91::ΔalmE | pWM91::ΔVC1579 | This study |
| Oligonucleotides | ||
| PcarRS-pVA289-F | AAACTCGAGAACACGCGGCGAGGAATTGAGTCAG | |
| PcarRS-pVA289-R | CGGGGATCCGATAATGTAGAGACTGGGTTGG | |
| P-vc1320-F-XhoI | ACCCTCGAGTAATCACTGAGAGTGTAGCC | |
| P-vc1320-R-XbaI | TTTTCTAGATGGGGACCTCGTATTTACGG | |
| P-vc1579-F-XhoI | AAACTCGAGATGTTGCGTCTATTGGCGCGCG | |
| P-vc1579-R-XbaI | AATTCTAGATCATGTCTTGATAGGTGT | |
| almE-F1 | CCCCCGGGCCACCAAGATACAAACTA | |
| almE-F2 | TACAATTCTGCGGCGAGTCAGACATA | |
| almE-R1 | ATGAGCTCGCTGCATCATGTCGGCTA | |
| almE-R2 | TGTCTGACTCGCCGCAGAATTGTATG | |
| carRS-F-EMSA | GCGGGAGTCGGCAGCGGGCGAGGAATTGAGTCAGAAGCC | |
| carRS-R-EMSA | GCGGGAGTCGGCAGCGGAGACTGGGTTGGTTAGACATGGGG | |
| 5′BIO | 5′-biotin-GCGGGAGTCGGCAGCG |