TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Genotype or descriptiona | Source or reference |
|---|---|---|
| E. coli strains | ||
| DH5α | supE44 ΔlacU169 (φ80 lacZΔM15) hsdR17 recA1 endA1 gyrA96 thi-1 relA1β | 18 |
| BL21(DE3) | hsdS gal (λcIts857 ind1 Sam7 nin5 lacUV5-T7 gene 1) | 28 |
| H. pylori strains | ||
| G27 | Clinical isolate; wild type | 32 |
| G27 fur (Δfur::Km) | G27 derivative with fur gene mutation; Kmr | 8 |
| G27 nikR (ΔnikR::Km) | G27 derivative with nikR gene mutation; Kmr | 23 |
| G27 fur nikR (Δfur::Km ΔnikR::cat) | G27 derivative with fur and nikR gene mutations; Kmr Cpr | 23 |
| G27 vac::PfecA3-lacZ | G27 derivative containing the wild-type PfecA3 promoter-lacZ fusion in the vacA locus; Kmr | This study |
| G27 vac::OP-I mutant PfecA3-lacZ | G27 derivative containing the OP-I mutant PfecA3 promoter-lacZ fusion in the vacA locus; Kmr | This study |
| G27 vac::OP-II mutant PfecA3-lacZ | G27 derivative containing the OP-II mutant PfecA3 promoter-lacZ fusion in the vacA locus; Kmr | This study |
| Plasmids | ||
| pGEM-T Easy | General cloning vector; Ampr | Promega |
| pGEMT-PfecA1 | pGEM-T Easy derivative containing 402 bp of fecA1 promoter region, amplified by PCR with primers A1F and A1R | This study |
| pGEMT-PfecA2 | pGEM-T Easy derivative containing 380 bp of fecA2 promoter region, amplified by PCR with primers A2F and A2R | This study |
| pGEMT-PfecA3 | pGEM-T Easy derivative containing 265 bp of fecA3 promoter region, amplified by PCR with primers A3F new and A3R new | This study |
| pBlueScript (pBS) | General cloning vector; Ampr | Stratagene |
| pBS-PfecA3Rx1 | pBlueScript derivative containing a 130-bp fragment of fecA3 promoter region, amplified by PCR with primers A3.1-A3.4 | This study |
| pBS-PfecA3Rx2 | pBlueScript derivative containing a 60-bp fragment of fecA3 promoter region, amplified by PCR with primers A3.1-A3.2 | This study |
| pBS-PfecA3Rx3 | pBlueScript derivative containing a 60-bp fragment of fecA3 promoter region, amplified by PCR with primers A3.3-A3.4 | This study |
| pBS-PA3MuOpI | pBlueScript derivative containing the 265-bp fecA3 promoter region with a SmaI site introduced into OP-I by PCR with the primers A3Fnew-A3.9 and A3Rnew-A3.8 | This study |
| pBS-PA3MuOpII | pBlueScript derivative containing the 265-bp fecA3 promoter region with a SmaI site introduced into OP-II by PCR with the primers A3Fnew-A3.7 and A3Rnew-A3.6 | This study |
| pVac::Km | pGEMZ derivative containing a kanamycin cassette | 10 |
| pVac::PA3lacZ | pVac::Km derivative containing the transcriptional fusion PfecA3-lacZ; Kmr | This study |
| pVac::PA3MuOpIlacZ | pVac::Km derivative containing the transcriptional fusion OP-I mutant PfecA3-lacZ; Kmr | This study |
| pVac::PA3MuOpIIlacZ | pVac::Km derivative containing the transcriptional fusion OP-II mutant PfecA3-lacZ; Kmr | This study |
| pET15b | IPTG-inducible vector overexpressing N-terminally His6-tagged recombinant protein; Ampr | Novagen |
| pET15b-nikR | pET15b derivative containing the nikR coding sequence cloned in frame within the NdeI/BamHI restriction sites | 7 |
| pET15b-fur | pET15b derivative containing the fur coding sequence cloned in frame within the NdeI/XhoI restriction sites | 8 |
a
IPTG, isopropyl-β-d-thiogalactopyranoside.