TABLE 1.
Plasmids and strains used in this study
| Plasmid or strain | Relevant characteristic(s)a | Source and/or reference |
|---|---|---|
| Plasmids | ||
| B. anthracis | ||
| pXO1 | Tox+ | 26 |
| E. coli | ||
| pBSIIKS+ | Apr | Stratagene |
| pG+host5 | Emr | Appligene |
| pGEM-T | Apr | Promega |
| pSL301 | Apr | Invitrogen |
| pUC18::Ωkm-2 | pUC18 carrying the Ωkm-2 cassette; Apr Kmr | 18 |
| pUTE40 | 3.0-kb HpaI-BamHI pXO1 fragment, containing pagA, in pMK4 | 12 |
| pUTE41 | 3.0-kb HpaI-BamHI pXO1 fragment, containing pagA, in pBSIIKS+ | This work |
| pUTE103 | 4.3-kb PstI-XbaI pXO1 fragment containing pagR and flanking sequences in pG+host5 | This work |
| pUTE308 | BamHI-KpnI PCR product containing 0.9 kb upstream of pagR and the 5′ end of pagR in pSL301 | This work |
| pUTE331 | 1.1-kb PCR product containing the 3′ end of pagA, the pagA attenuator, and the 5′ end of pagR in pGEM-T | This work |
| pUTE333 | 2.6-kb BamHI-SacI fragment from pUTE103, containing pagR, in pUTE41 | This work |
| Bifunctional | ||
| pUTE29 | Apr in E. coli, Tcr in B. anthracis | 12 |
| pUTE34 | 2.7-kb SnaBI-EcoRI pXO1 fragment containing atxA in pUTE29 | 12 |
| pUTE314 | 4.0-kb SacI-PstI pXO1 fragment containing pagR and flanking sequences in pUTE29 | This work |
| pUTE315 | Ωkm-2 cloned into BamHI site between pagA and pagR in pUTE314 | This work |
| pUTE334 | 5.6-kb SacI-KpnI fragment from pUTE333, containing pagA and pagR, in pUTE29 | This work |
| pUTE366 | pUTE334 containing +1 frameshift within pagR | This work |
| Strains | ||
| B. anthracis | ||
| UM44b | Tox+ Ind− | 26 |
| UT53 | atxA-Null derivative of UM44, atxA is replaced by Ωkm-2; Ind− Kmr | 7 |
| UT62 | P1 start site of pagA disrupted with the Ωkm-2, derivative of UM44; Ind− Kmr | 7 |
| UT82 | Tn917-LTV3 insertion mutant of UM44; Cmr MLSr, CO2-atxA-regulated lacZ expression | 10 |
| UT119 | pagR Mutant derivative of UM44, Ωkm-2 inserted at BamHI site between pagA and pagR; Ind− Kmr | This work |
| E. coli | ||
| GM1684 | F′ F-lacIqΔM15 pro+/dam-4 Δ(lac-pro)X111 thi-1 glnV44 (relA1) | R. Kolter |
| JM109 | F′ traD36 proA+ proB+ lacIqlacZΔM15/recA1 endA1 gyrA96 thi hsdR17 supE44 relA1 Δ(lac-proAB) mcrA | 35 |
| BMH 71-18 | thi supE Δ(lac-proAB) [mutS::Tn10][F′ proAB lacIqZΔM15] | Clontech |
a
Abbreviations: Apr, ampicillin resistant; Emr, erythromycin resistant; Ind, indole; Kmr, kanamycin resistant; MLSr, macrolide resistant; Tcr, tetracycline resistant; Tox, anthrax toxin proteins.
b
UM44 was derived from the Weybridge strain.