Table 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant genotype or descriptiona | Reference or source |
|---|---|---|
| E. coli strains | ||
| MG1655 | F− λ− ilvG rfb50 rph-1 | 8 |
| MG1655 rspL150 | F− λ− ilvG rfb50 rph-1 rpsL150; Strr | 26 |
| MG2 | MG1655 ΔyehUT | This work |
| MG3 | MG1655 rpsL150 ΔyehT∷rpsL-neo; Kanr Strs | This work |
| MG6 | MG1655 rpsL150 ΔyehU∷rpsL-neo; Kanr Strs | This work |
| MG10 | MG1655 ΔyjiY∷FRT-PGK-gb2-neo-FRT; Kanr | This work |
| MG11 | MG1655 ΔcyaA∷FRT-PGK-gb2-neo-FRT; Kanr | This work |
| MG12 | MG1655 Δcrp∷FRT-PGK-gb2-neo-FRT; Kanr | This work |
| MG1655-ΔlacZ | MG1655 ΔlacZ∷Tetr | K. Jahreis (personal gift) |
| BL21(DE3) | F− ompT hsdSB(rB− mB−) gal dcm (DE3) | 58 |
| Plasmids | ||
| pRed/ET | λ RED recombinase in pBAD24; Ampr | Gene Bridges |
| pCP20 | FLP recombinase, λcI 857+, λpR Repts; Ampr Cmr | 15 |
| pBAD18-Kan | Arabinose-inducible PBAD promoter, pBR322 ori; Kanr | 24 |
| pBAD18-yehT | 6His-yehT and Shine-Dalgarno sequence cloned into the NheI and SphI sites of pBAD18-Kan; Kanr | This work |
| pBAD24 | Arabinose-inducible PBAD promoter, pBR322 ori; Ampr | 24 |
| pBAD24-yehT | 6His-yehT cloned into the EcoRI and XbaI sites of pBAD24; Ampr | This work |
| pBAD24-yehT D54E | yehT D54E cloned into the EcoI and XbaI sites of pBAD24-yehT; Ampr | This work |
| pBAD24-yehT D54N | yehT D54N cloned into the NdeI and XbaI sites of pBAD24-yehT; Ampr | This work |
| pBAD24-kdpE | kdpE cloned into the NdeI and XbaI sites of pBAD24-kdpE; Ampr | This work |
| pBAD24-yjiY | yjiY-6His cloned into the EcoRI and XbaI sites of pBAD24; Ampr | This work |
| pBAD24-yehU | yehU-6His cloned into the EcoRI and XbaI sites of pBAD24; Ampr | This work |
| pBAD24-yehU H382Q | yehU H382Q cloned into the EcoRI and NdeI sites of pBAD24-yehU; Ampr | This work |
| pBAD24-yehUT | yehUT cloned into the EcoRI and XbaI sites of pBAD24; Ampr | This work |
| pUC19 | IPTG-inducible Plac promoter, pMB1 ori; Ampr | 66 |
| pUC19 PyjiY−212/+88 | PyjiY−212/+88 cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY−12/+88 | PyjiY−12/+88 cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY−112/−13 | PyjiY −112/−13 cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY−212/−113 | PyjiY−212/−113 cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY MM3 | PyjiY MM3 (substituted M3 site) cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY MM23 | PyjiY MM23 (substituted M2 or M3 site) cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY MM123 | PyjiY MM123 (substituted M1 to M3 sites) cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pUC19 PyjiY SC14 | PyjiY SC14 (substituted spacer motif, 14 nucleotides replaced) cloned into the EcoRI and BamHI sites of pUC19; Ampr | This work |
| pRS415 | Operon fusion vector | 55 |
| pRS415 PyjiY−212/+88 | PyjiY−212/+88 cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY MM3 | PyjiY MM3 (substituted M3 site) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY MM23 | PyjiY MM23 (substituted M2 and M3 sites) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY MM123 | PyjiY MM123 (substituted M1 to M3 sites) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY SC14 | PyjiY SC14 (substituted spacer motif, 14 nucleotides replaced) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY SC1 | PyjiY SC1 (central nucleotide within the 13-bp spacer replaced) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY SC3 | PyjiY SC3 (three central nucleotides within the 13-bp spacer replaced) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY SC7 | PyjiY SC7 (seven central nucleotides within the 13-bp spacer replaced) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pRS415 PyjiY SC13 | PyjiY SC13 (13 nucleotides within the 13-bp spacer replaced) cloned into the EcoRI and BamHI sites of pRS415; Ampr | This work |
| pBBR1-MCS5-TT-RBS-lux | luxCDABE and terminators lambda T0 rrnB1 T1 cloned into pBBR1-MCS5 for plasmid-based transcriptional fusions; Gmr | 23 |
| pBBR yjiY-lux | PyjiY −212/+88 cloned into the BamHI and EcoRI sites of pBBR1-MCS5-TT-RBS-lux; Gmr | This work |
| pBBR yjiY-sub-CRP-BS-lux | PyjiY sub-CRP-BS (substituted CRP-binding site in PyjiY−212/+88) cloned into the BamHI and EcoRI sites of pBBR1-MCS5-TT-RBS-lux; Gmr | This work |
a
IPTG, isopropyl-β-d-thiogalactopyranoside.