TABLE 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant characteristicsa | Reference or source |
|---|---|---|
| E. coli | ||
| DH5α λpir | pir lysogen Δ(ara-leu) araD Δ(lacX74) phoA20 thi-1 rpoB argE(Am) recA1 | 40 |
| SM10 λpir | thi thir leu tonA lecY supE recA::RP4-2-Tc::Mu Kmr (pirR6K) | 41 |
| BL21(DE3) | Host strain for protein expression | Novagen |
| V. alginolyticus | ||
| EPGS | Wild-type pathogenic isolate from fish; Ampr | 25 |
| XT001 | EPGS asp−; Cmr | 16 |
| XT002 | EPGS ΔluxR | 16 |
| XT003 | EPGS ΔaphA | 20 |
| XT004 | EPGS ΔaphB | This study |
| XT005 | EPGS ΔaphA ΔaphB | This study |
| XT020 | ΔaphB/pBAD33-aphB-flag; Cmr | This study |
| XT021 | aphB+, ΔaphB/pBAD33-aphB-flag2; Cmr | This study |
| XT022 | ΔaphB/pBAD33-aphB; Cmr | This study |
| XT023 | ΔaphB/pBAD33; Cmr | This study |
| XT024 | EPGS/pBAD33-Flag; Cmr | This study |
| XT025 | EPGS/pDM8-Pasp-luxAB; Cmr | This study |
| XT026 | ΔaphB/pDM8-Pasp-luxAB; Cmr | This study |
| XT027 | EPGS/pDM8-PaspMBSI/II-luxAB; Cmr | This study |
| XT028 | EPGS/pDM8-PluxR-luxAB; Cmr | This study |
| XT029 | ΔaphB/pDM8-PluxR-luxAB; Cmr | This study |
| XT030 | ΔluxR/pDM8-PluxR-luxAB; Cmr | This study |
| XT031 | ΔaphA/pDM8-PluxR-luxAB; Cmr | This study |
| XT032 | ΔaphA ΔaphB/pDM8-PluxR-luxAB; Cmr | This study |
| XT034 | EPGS/pDM8-PaphB-luxAB; Cmr | This study |
| XT035 | ΔaphB/pDM8-PaphB-luxAB; Cmr | This study |
| XT036 | EPGS/pDM8-PluxRΔBBS-luxAB; Cmr | This study |
| XT037 | EPGS/pDM8-PluxRΔABS-luxAB; Cmr | This study |
| XT038 | EPGS/pDM8-PluxRΔRBSI-luxAB; Cmr | This study |
| XT040 | EPGS/pDM8-PluxRMBBS-luxAB; Cmr | This study |
| XT041 | EPGS/pDM8-PluxRMRBSI-luxAB; Cmr | This study |
| XT042 | EPGS/pDM8-PluxRMRBSPI-luxAB; Cmr | This study |
| XT043 | EPGS/pDM8-PluxRΔABSMBBS-luxAB; Cmr | This study |
| XT057 | BL21/pET28a-aphA; Kmr | 20 |
| XT058 | BL21/pET28a-luxR; Kmr | 16 |
| XT046 | BL21/pET28a-aphB; Kmr | This study |
| XT050 | PaspMBBSI/II, Pasp with scrambled AphB binding sites I/II | This study |
| XT051 | EPGS/pDM8-PaspMBBSI-luxAB; Cmr | This study |
| XT052 | EPGS/pDM8-PaspMBBSII-luxAB; Cmr | This study |
| XT053 | ΔaphB/pDM8-PaspMBBSI-luxAB; Cmr | This study |
| XT054 | ΔaphB/pDM8-PaspMBBSII-luxAB; Cmr | This study |
| XT055 | ΔaphB/pDM8-PaspMBBSI/II-luxAB; Cmr | This study |
| Plasmids | ||
| pDM4 | Suicide vector, pir dependent, R6K, SacBR; Cmr | 42 |
| pBAD33-mob | pBAD33, mob+ allowing for conjugation; Cmr | 20 |
| pDM8 | pSup202 containing promoterless lacZ; Cmr | 43 |
| pET28a | Expression vector; Kmr | Novagen |
| pDM4::aphA | pDM4 with aphA fragment deletion of 4 to 573 bp | 20 |
| pDM4-aphB | pDM4 carrying aphB with deletion of 4 to 873 bp | This study |
| pBAD33-aphB-flag | pBAD33-derived aphB-Flag expression plasmid | This study |
| pBAD33-aphB-flag2 | pBAD33 derivative expressing aphB-Flag by PaphB | This study |
| pBAD33-flag | pBAD33 derivative expressing Flag alone | This study |
| pBAD33-aphB | pBAD33-derived aphB expression plasmid | This study |
| pET28a-aphB | pET28a with aphB ORF | This study |
| pET28a-aphA | pET28a with aphA ORF | 20 |
| pET28a-luxR | pET28a with luxR ORF | 18 |
| pDM8-Pasp-luxAB | pDM8 with Pasp-luxAB | This study |
| pDM8-PaspMBBSI/II-luxAB | pDM8 with mutated AphB boxes of Pasp-luxAB | This study |
| pDM8-PaspMBBSI-luxAB | pDM8 with mutated AphB box I of Pasp-luxAB | This study |
| pDM8-PaspMBBSII-luxAB | pDM8 with mutated AphB box II of Pasp-luxAB | This study |
| pDM8-PluxR-luxAB | pDM8 with PluxR-luxAB | This study |
| pDM8-PaphB-luxAB | pDM8 with PaphB-luxAB | This study |
| pDM8-PluxRΔRBSI-luxAB | pDM8-PluxR-luxAB with LuxRI box deleted | This study |
| pDM8-PluxRΔABS-luxAB | pDM8-PluxR-luxAB with AphA box deleted | This study |
| pDM8-PluxRΔBBS-luxAB | pDM8-PluxR-luxAB with AphB box deleted | This study |
| pDM8-PluxRMBBS-luxAB | pDM8-PluxR-luxAB with AphB box mutated | This study |
| pDM8-PluxRMRBSI-luxAB | pDM8-PluxR-luxAB with LuxRI box mutated | This study |
| pDM8-PluxRMRBSPI-luxAB | pDM8-PluxR-luxAB with LuxRI box partially mutated | This study |
| pDM8-PluxRΔABSMBBS-luxAB | pDM8-PluxR-luxAB with AphB box mutated and AphA box deleted | This study |
a
Kmr, kanamycin resistant; Ampr, ampicillin resistant; Cmr, chloramphenicol resistant.