TABLE 1.
E. coli strains and plasmid constructs
| Strain or plasmid | Relevant characteristic(s) | Relevant genotype | Source or reference |
|---|---|---|---|
| Strains | |||
| DH5α | Standard host strain for cloning | φ80dlacZΔM15 Δ(lacZYA-argF) endA1 recA1 | Gibco-BRL |
| AG102 | Derived from AG100 | marR1 | 9 |
| AG102A | Derived from AG102 | marR1 ΔacrAB::kan | 27 |
| AG102MB | Derived from AG102 | marR1 actB::kan | 7 |
| HNCE1a | Derived from AG102MB | marR1 acrB::kan ΔacrD | 7 |
| HNCE1b | Derived from HNCE1a | marR1 acrB::kan ΔacrD acrA::cat | 7 |
| HNCE3 | Derived from AG102 | marR1 ΔacrA | This study |
| HNCE5 | Derived from AG102A | marR1 ΔacrAB::kan ΔyhiV::cat | This study |
| Plasmidsa | |||
| pKD46 | λ red recombinase expression plasmid; ara-inducible expression, temperature-sensitive replication | amp λ(γ, β, exo) araC | 5 |
| pKD3 | Template plasmid; FRT-flanked cat | amp cat oriRγ | 5 |
| pCP20 | FLP expression plasmid; temperature-sensitive replication and FLP synthesis | amp flp | 5 |
| pSportIb | High-copy-number cloning and expression vector; lac-inducible expression | amp lacI | Gibco-BRL |
| pAcrBc | Clone from DH5α | acrB | 7 |
| pAcrDc | Clone from DH5α | acrD | 7 |
| pYhiVc | Clone from DH5α | yhiV | This study |
| pMdtBCc | Clone from DH5α | mdtB mdtC | This study |
| pAcrAc | Clone from DH5α | acrA | This study |
| pAcrEc | Clone from DH5α | acrE | This study |
| pYhiUc | Clone from DH5α | yhiV | This study |
| pMdtAc | Clone from DH5α | mdtA | This study |
a
See Fig. 3 for plasmids expressing AcrA-YhiU chimeras.
b
Plasmid vector used for all constructs created in this study.
c
The lac-inducible promoter of pSportI controls the expression of all cloned gene constructs created in this study.