Unraveling CRP/cAMP-mediated metabolic regulation in Escherichia coli persister cells

  1. Han G Ngo
  2. Sayed Golam Mohiuddin
  3. Aina Ananda
  4. Mehmet Orman  Is a corresponding author
  1. Department of Chemical and Biomolecular Engineering, University of Houston, United States
  2. Department of Biology, Monmouth University, United States
7 figures, 1 table and 8 additional files

Figures

Figure 1 with 8 supplements
Crp/cAMP regulation of persister cell formation in the stationary phase.

E. coli K-12 MG1655 WT and mutant cells at early (t=5 hr) and late (t=24 hr) stationary phases were transferred to fresh medium with antibiotics for persister cell quantification. At time points 0, 2, 4, 6, 8, and 20 hr, 1 mL of the treated culture was washed with 1 X phosphate-buffered saline (PBS) to remove antibiotics. It was then serially diluted and plated on an agar plate to count the colony-forming units (CFUs). (A) Persister levels of ampicillin-treated culture with an antibiotic concentration of 200 μg/mL. (B) Persister levels of ofloxacin-treated culture with an antibiotic concentration of 5 μg/mL. (C) Persister levels of gentamicin-treated culture with an antibiotic concentration of 50 μg/mL. The number of biological replicates is n=4 for all panels. Biphasic kill curves were generated using a non-linear model (see Materials and methods). Statistical significance tests were conducted using F-statistics (*p < 0.05, **p < 0.01, ****p < 0.0001). The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 1
cAMP concentrations normalized to cell numbers for E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA.

The cAMP levels were measured in late stationary phase cultures at 450 nm using the Cyclic AMP XP Assay Kit (Cell Signaling Technology). n=4. Statistical significance was observed between control and mutant strains (*p<0.05, ***p<0.001, One-way ANOVA with Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 2
Genetic perturbation of Crp/cAMP enhanced PcyaA promoter activity, resulting in increased gfp expression.

Overnight cultures of E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA strains harboring the pMSs201 plasmid, which encodes green fluorescent protein (GFP) under the control of the PcyaA promoter, were diluted 1:1000 into fresh LB medium and incubated at 37 °C with shaking at 250 rpm for 24 hr. Cells at the late stationary phase were then collected, diluted in 1 X PBS, and analyzed by flow cytometry. n=4. Statistical significance was observed between control and mutant strains (****p<0.0001, One-way ANOVA with Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 3
Growth curves of E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA.

Optical densities of cell cultures at 600 nm (OD600) were measured every hour using a plate reader. n=3. The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 4
Normalized cAMP concentrations of E. coli K-12 MG1655 WT.

The cAMP concentrations were measured in growth cultures at the indicated time points. First, the cAMP concentrations were normalized to the number of cells. Subsequently, the data were further normalized based on the time point 0 to mitigate errors associated with batch-to-batch assay kit variations and to capture the trend in cAMP levels across the time points. n=8. Statistical significance was observed between time points (**p<0.01, One-way ANOVA with Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard error.

Figure 1—figure supplement 5
Persister levels of cells carrying the Crp expression system.

Cells at early (t=5 hr) and late (t=24 hr) stationary phases were transferred to fresh media with antibiotics for persister cell quantification. At time points 0, 2, 4, 6, 8, and 20 hr, 1 mL of the treated culture underwent two washes with 1 X PBS to remove antibiotics. It was then serially diluted and plated on an agar plate to count the CFUs. (A) Persister levels of ampicillin-treated culture (200 μg/mL). (B) Persister levels of ofloxacin-treated cultures (5 μg/mL). (C) Persister levels of gentamicin-treated culture (50 μg/mL). n=4. Biphasic kill curves were generated using a non-linear model (see Materials and methods). Statistical significance tests were conducted using F-statistics (**p < 0.01 and ****p < 0.0001). The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 6
Persister levels of E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA strains at normalized antibiotic concentrations.

Panels show survival following treatment with (A) ampicillin, (B) ofloxacin, and (C) gentamicin. After treatment, samples were washed six times with 1 X PBS to minimize antibiotic carryover. Antibiotic concentrations were normalized to MICs to ensure valid comparisons across strains (The concentrations of 33×MIC for ampicillin and 100×MIC for ofloxacin and gentamicin match those used in Figure 1). n=4. Statistical significance was observed between control and mutant strains (****p<0.0001, One-way ANOVA with Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 1—figure supplement 7
Agar plates showing E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA strains following treatment with antibiotics at normalized concentrations.

After treatment, cells were washed and subjected to 10-fold serial dilutions, then plated on agar. Plates were incubated for (A) 16  hr, (B) 48  hr, and (C) 72  hr to assess colony formation over time. The panel is a representative biological replicate. Consistent results were seen across all three biological replicates.

Figure 1—figure supplement 8
Deletion of crp and cyaA reduces ampicillin and ofloxacin persistence in the hipA7 strain.

Persistence levels were assessed by exposing E. coli K-12 MG1655 WT, hipA7, hipA7Δcrp, and hipA7ΔcyaA strains in the late stationary phase to the specified antibiotics, followed by CFU quantification at designated time points. (A) Ampicillin (200 µg/ml), (B) Ofloxacin (5 µg/ml) and (C) Gentamicin (50 µg/ml). n=4. Statistical significance was observed between control and mutant strains (****p<0.0001, One-way ANOVA with Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 2 with 3 supplements
The effect of Crp/cAMP on persister cell metabolism during stationary phase.

(A) MS analysis of E. coli K-12 MG1655 WT, and Δcrp at early (t=5 hr) and late (t=24 hr) stationary phases. Unsupervised hierarchical clustering was applied to standardized metabolic data. Each column represents a biological replicate. n=4. (B) Pathway enrichment analysis was conducted using MetaboAnalyst (Lu et al., 2023). Upregulated and downregulated pathways of the Δcrp strain compared to WT in the late stationary growth phase were provided in this figure. (C, D) Pathway enrichment maps comparing metabolites of the TCA cycle, pentose phosphate metabolism, glycolysis, gluconeogenesis, and pyruvate metabolism in Δcrp versus WT for early and late stationary phase conditions, respectively. Circle size corresponds to the ratio of normalized metabolite intensities between mutant and control cells. Blue (p ≤ 0.05 for dark blue; 0.05 < p  <  0.10 for light blue) and red (p ≤ 0.05 for dark red; 0.05 < p  <  0.10 for light red) indicate significantly downregulated or upregulated metabolites in the mutant compared to the control. White signifies no significant difference. n=4 for all panels. ESP: Early stationary phase, LSP: Late stationary phase.

Figure 2—source data 1

Normalized metabolomics data of early and late stationary phases of wild-type and mutant crp across three biological replicates.

Data collected and analyzed by Metabolon Inc (Morrisville, NC).

https://cdn.elifesciences.org/articles/99735/elife-99735-fig2-data1-v1.xlsx
Figure 2—figure supplement 1
The pathway enrichment analysis comparing WT and Δcrp strains during the early stationary phase cultures.

The analysis was conducted using MetaboAnalyst, with a threshold ratio (Δcrp/WT) set at ≤0.5 for downregulation and ≥2 for upregulation. (A) Upregulated and (B) Downregulated pathways in the Δcrp strain compared to WT during the early stationary growth phase (ESP).

Figure 2—figure supplement 2
The pathway enrichment analysis for the WT strain.

The analysis was conducted using MetaboAnalyst, with a threshold ratio (LSP/ESP) set at ≤0.5 for downregulation and ≥2 for upregulation. (A) Upregulated and (B) Downregulated pathways of the WT strain in the late stationary growth phase (LSP) compared to the WT strain in the early stationary phase (ESP).

Figure 2—figure supplement 3
The pathway enrichment analysis for the Δcrp strain.

The analysis was conducted using MetaboAnalyst, with a threshold ratio (LSP/ESP) set at ≤0.5 for downregulation and ≥2 for upregulation. (A) Upregulated and (B) Downregulated pathways of the Δcrp strain in the late stationary growth phase (LSP) compared to the Δcrp strain in the early stationary phase (ESP).

Figure 3 with 1 supplement
Validation of Crp/cAMP-mediated metabolic state in persister cells through proteomics analysis.

Pathway enrichment analysis was conducted in STRING (Szklarczyk et al., 2021; Szklarczyk et al., 2023) for upregulated (A) and downregulated (B) proteins. Genes highlighted in red are linked with the upregulated protein networks, while genes in blue, gray, and purple correspond to those in the downregulated protein network. The visual network in STRING illustrates protein interactions. In evidence mode, color in the network represents the interaction evidence of data support, derived from curated databases, experimental data, gene neighborhood, gene fusions, co-occurrence, co-expression, protein homology, and text mining (Szklarczyk et al., 2021; Szklarczyk et al., 2023).

Figure 3—source data 1

Normalized proteomics data of the late stationary phase of wild-type and mutant crp across three biological replicates.

Data collected by UT Health’s Clinical and Translational Proteomics Service Center (Houston, TX).

https://cdn.elifesciences.org/articles/99735/elife-99735-fig3-data1-v1.xlsx
Figure 3—figure supplement 1
The MS analysis of proteins from both WT and Δcrp strains at the late stationary phase.

The proteomic data were subjected to unsupervised hierarchical clustering. Each column in the figure represents a biological replica. n=3.

Figure 4 with 2 supplements
The role of Crp/cAMP in non-growing cell formation.

(A, B) Flow cytometry histograms depict mCherry expression in E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA at early (t=5 hr) and late (t=24 hr) stationary phases, respectively. Cells containing an IPTG-inducible mCherry expression system were cultivated with IPTG. After washing and dilution of early and late stationary phase cells in IPTG-free fresh media, fluorescence was tracked in non-growing and growing cells for 2.5 hr. The panel is a representative biological replicate. Consistent results were seen across all three biological replicates. (C) Growth curves of WT, Δcrp, and ΔcyaA cultures were determined using flow cytometry to calculate lag and doubling times. Lag times were calculated using the ‘Microbial lag phase duration calculator’ (Opalek et al., 2022). Doubling times were computed using the formula td=Δt/(3.3xLog10(N/No)). n=3. *Statistical significance observed between control and mutant strains (p<0.05, two-tailed t-test). The data for each time point represent the mean value  ± standard deviation.

Figure 4—figure supplement 1
Persister levels of E. coli WT, Δcrp, and ΔcyaA cells with the integrated mCherry expression system.

Late stationary phase cultures (t=24 hr) were transferred to fresh media and treated with ampicillin (200 μg/mL), ofloxacin (5 μg/mL), and gentamicin (50 μg/mL) for 20 hr. Subsequently, 1 mL of the treated culture underwent two washes with 1 X PBS to remove antibiotics. It was then serially diluted and plated on an agar plate to count the CFUs. The levels of ofloxacin and gentamicin persisters in the mutant strains were below the limit of detection. n=4. Statistical significance was observed between control and mutant strains (***p<0.001, ****p<0.0001, Two-way ANOVA with Tukey’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 4—figure supplement 2
Non-growing cell levels in the E. coli strain carrying the Crp expression system.

(A, B) Flow cytometry histograms depict mCherry expression in Δcrp +pUA66 crp, and Δcrp +pUA66 EV at early (t=5 hr) and late (t=24 hr) stationary phases, respectively. Cells containing an IPTG-inducible mCherry expression system were cultivated with IPTG. After washing and dilution of early and late stationary phase cells in IPTG-free fresh media, fluorescence was tracked in non-growing and growing cells for 2.5 hr. The panel is a representative biological replicate. Consistent results were seen across all three biological replicates.

Figure 5 with 4 supplements
Crp/cAMP-mediated metabolic state of persister cells.

(A) GFP reporter plasmid introduced into E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA cells to monitor SQR gene activity. Flow cytometry was used to detect activity at early (t=5 hr) and late (t=24 hr) stationary phases. The panel on the left represents a biological replicate, and the results are consistent across all three replicates, as demonstrated in the panel on the right. Statistical significance observed between control and mutant groups (*p<0.05, **p<0.01, ***p<0.001, two-tailed t-test). (B) Redox activities of E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA cells were measured at early (t=5 hr) and late (t=24 hr) stationary phases by flow cytometry using a RSG dye. This dye fluoresces green after reduction by bacterial reductases. A representative biological replicate is shown (left), with consistent results across all five replicates (right). Statistical significance observed between control and mutant groups (*p<0.05, **p<0.01, two-tailed t-test). (C) E. coli cells with integrated mCherry expression system used to validate cellular respiration. Cells were diluted into fresh media and treated with ampicillin (200 μg/mL) for 20 hr. Flow cytometry measured the red fluorescence of intact surviving cells. A representative biological replicate is shown, with consistent results across all three replicates. (D) RSG levels of cells (carrying the mCherry expression system) at exponential phase (t=3 hr); cells before ampicillin treatment; non-lysed (intact) cells after 20 hr of ampicillin treatment; and untreated cells after 20 hr of culturing. A representative biological replicate is shown (left), with consistent results across all four replicates (right). Statistical significance observed between intact antibiotic-treated cells and others (*p<0.05, **p<0.01, two-tailed t-test). (E) High-throughput screening of mutants from the Keio collection. The mutant strains selected are associated with central metabolism. Stationary phase cells were diluted 100-fold in fresh medium and treated with ampicillin (200 μg/mL) or ofloxacin (5 μg/mL) for 20 hr. Treated cultures were washed, serially diluted, and plated on agar plates to quantify CFUs. (F) Genes related to the TCA cycle, ETC, ATP synthase, glycolysis, and pentose phosphate pathway (PPP) were knocked out and then treated with ampicillin (200 μg/mL) or ofloxacin (5 μg/mL) to enumerate CFUs. n=4. Biphasic kill curves were generated using a non-linear model. Statistical significance tests were conducted using F-statistics (*p < 0.05, and **p < 0.01). Each data point represents the mean value  ± standard deviation.

Figure 5—figure supplement 1
RSG staining control for bacterial metabolic activities.

Exponential phase (t=3 hr) cells were stained with 1 μM RSG for 10 min at 37 °C before analyzing by flow cytometry. Unstained cells and cells treated with 20 μM CCCP +1 μM RSG were used as control. CCCP was expected to reduce cellular redox activities. The panel is a representative biological replicate. Consistent results were seen across all three biological replicates.

Figure 5—figure supplement 2
Intact (non-lysed) cell levels of E. coli WT, Δcrp, and ΔcyaA cells with the integrated mCherry expression system.

mCherry-positive cells were diluted into fresh media and treated with ampicillin (200 μg/mL) for 20 hr. Flow cytometry was used to measure the red fluorescence of intact surviving cells at time points t=0, 1, 2, 4, 6, and 20 hr. A representative biological replicate is shown, with consistent results across all three replicates.

Figure 5—figure supplement 3
VBNC levels of E. coli WT, Δcrp, and ΔcyaA cells with the integrated mCherry expression system.

Flow cytometry was employed to quantify intact surviving cells. Persister cells were quantified by plating the cells on agar media. Viable but nonculturable (VBNC) cells were enumerated by subtracting persister levels from the intact cell levels. n=4. Statistical significance was observed between control and mutant strains (****p<0.0001, One-way ANOVA using Dunnett’s multiple comparisons test). The data for each time point represent the mean value  ± standard deviation.

Figure 5—figure supplement 4
Cell counts of E. coli K-12 MG1655 WT and mutant strains using flow cytometry at late stationary phase.

Cells were diluted 100-fold into 1 mL of 1 X PBS. n=4. The data for each time point represent the mean value  ± standard deviation.

Author response image 1
Persister levels of E. coli K-12 MG1655 WT, Δcrp, and Δcrp strains in late stationary phase.

Cells were treated with ampicillin (5× MIC for 4 h), ofloxacin (5× MIC for 2.5 h), and gentamicin (3× MIC for 1 h). Concentrations and treatment durations were selected based on (Zeng et al., 2022).

Author response image 2
Persister levels of E. coli K-12 MG1655 (Panel A) and BW25113 (Panel B) WT, Δcrp, and Δcrp strains in the exponential growth phase.

Cells were treated at mid-exponential phase (OD600 ~0.25) with ampicillin (5× MIC for 4 h), ofloxacin (5× MIC for 2.5 h), and gentamicin (3× MIC for 1 h). Treatment concentrations and durations were based on conditions described in (Zeng et al., 2022).

Tables

Appendix 1—key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
Strain, strain background (Escherichia coli)K-12 MG1655
Wild Type
Gift from Dr. Mark P. Brynildsen
Strain, strain background (E. coli)K-12 MG1655
hipA7
Gift from Dr. Mark P. Brynildsen
Strain, strain background (E. coli)K-12 MG1655
hipA7Δcrp
This study
Strain, strain background (E. coli)K-12 MG1655
hipA7ΔcyaA
This study
Strain, strain background (E. coli)K-12 BW25113
Wild Type
Keio collectionCatalog # OEC5042
Strain, strain background (E. coli)K-12 MG1655
MO
Gift from Dr. Mark P. Brynildsen
Strain, strain background (E. coli)K-12 MG1655
MO Δcrp
This study
Strain, strain background (E. coli)K-12 MG1655
MO ΔcyaA
This study
Strain, strain background (E. coli)K-12 MG1655
Δcrp
This study
Strain, strain background (E. coli)K-12 MG1655
ΔcyaA
This study
Strain, strain background (E. coli)K-12 MG1655
ΔsucA
This study
Strain, strain background (E. coli)K-12 MG1655
Δlpd
This study
Strain, strain background (E. coli)K-12 MG1655
ΔsucC
This study
Strain, strain background (E. coli)K-12 MG1655
ΔsdhA
This study
Strain, strain background (E. coli)K-12 MG1655
ΔgltA
This study
Strain, strain background (E. coli)K-12 MG1655
ΔaceE
This study
Strain, strain background (E. coli)K-12 MG1655
ΔtktB
This study
Strain, strain background (E. coli)K-12 MG1655
Δmdh
This study
Strain, strain background (E. coli)K-12 MG1655
ΔnuoI
This study
Strain, strain background (E. coli)K-12 MG1655
ΔnuoM
This study
Strain, strain background (E. coli)K-12 MG1655
ΔatpA
This study
Strain, strain background (E. coli)K-12 MG1655
ΔatpB
This study
Strain, strain background (E. coli)K-12 MG1655
ΔatpC
This study
Strain, strain background (E. coli)K-12 MG1655
ΔatpD
This study
Strain, strain background (E. coli)K-12 MG1655
ΔfrdC
This study
Strain, strain background (E. coli)K-12 MG1655
Δpgi
This study
Strain, strain background (E. coli)K-12 MG1655
Δzwf
This study
Strain, strain background (E. coli)K-12 MG1655
ΔtalA
This study
Strain, strain background (E. coli)K-12 BW25113
ΔacnB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsucA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsucB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsucC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsdhD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsdhC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsdhB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsdhA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔaceF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtalB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcyoA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcyoB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcyoC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcyoD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔacnA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δicd
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfumA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfumB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfumC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δmdh
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δpgi
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpfkA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtpiA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔgpmM
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔppsA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpykF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpykA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔmaeB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δpck
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔrpiB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δrpe
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtktB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtalA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δzwf
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δgnd
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δppc
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfrdA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfrdB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfrdC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfrdD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔadhE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpflB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔaceB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔaceA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglcB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoJ
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoK
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoL
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoM
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoN
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔappC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔappB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔgltA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δlpd
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δfbp
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔaceE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtktA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δpta
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔackA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔldhA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δdld
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcydB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpoxB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglpX
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔybhA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔcydX
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfumE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔyggF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δccp
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔyieF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔwrbA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdhF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnrfD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnrfC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnrfA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔputA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔdmsC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtorC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtorA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhyaA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhyaB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhyaC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔkefF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarV
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔkduI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔeutE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhycE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhycG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δedd
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δeda
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdnG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdnI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglpD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglpA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglpB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔglpC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhybO
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhybC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarY
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarZ
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnapG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnapH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnapA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpurT
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdoG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdoI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpF
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔphoA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔadhP
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔeutD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔdmsA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhybB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnapB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpI
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔmaeA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔpfkB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfbaB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔrpiA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfumD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔsucD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdnH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔdmsB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
Δndh
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarG
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnarH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔtdcE
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhycB
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhycC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhycD
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔfdoH
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔhybA
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔnuoC
Keio collectionCatalog # OEC4988
Strain, strain background (E. coli)K-12 BW25113
ΔatpG
Keio collectionCatalog # OEC4988
Recombinant DNA reagentpMSs201 (kanR)Dharmacon Promoter LibraryCatalog # OEC4988
Recombinant DNA reagentpUA66-EV (empty vector)Gift from Dr. Mark P. BrynildsenA DNA fragment including T5 promoter, KanR gene, pUA66 origin of replication and lacIq was amplified from the pUA66-gfp plasmid with primers having BspHI cut sites. The amplified DNA fragment was digested with BspHI, and then self-ligated to obtain the modified pUA66-EV that does not have the gfp gene.
Recombinant DNA reagentpUA66-crpThis studyThe crp gene with its promoter was amplified from the genomic DNA of E. coli, using forward and reverse primers with BglII and ScaI restriction enzyme cut sites, respectively. The pUA66-gfp plasmid was double digested with BglII and ScaI to remove T5 promoter region and gfp gene. Then, the digested crp gene with its promoter and plasmid were ligated to generate pUA66-crp.
Strain, strain background (E. coli)K-12 MG1655
pMSs201 PsdhABCD-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
Δcrp pMSs201 PsdhABCD-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
ΔcyaA pMSs201 PsdhABCD-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
pMSs201 PcyaA-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
Δcrp pMSs201 PcyaA-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
ΔcyaA pMSs201 PcyaA-gfp
This study
Strain, strain background (E. coli)K-12 MG1655
Δcrp pUA66-EV
This study
Strain, strain background (E. coli)K-12 MG1655
Δcrp pUA66-crp
This study
Software, algorithmPrism (version 10.3.0)GraphPadRRID:SCR_002798http://www.graphpad.com/
Software, algorithmFlowJo (version 10.8.1)Becton, Dickinson & CompanyRRID:SCR_008520https://www.flowjo.com/
Software, algorithmMATLAB (version R2020b)MathWorksRRID:SCR_001622https://www.mathworks.com/
Sequence-based reagentForward Primer (5’ to 3’)
Δcrp::KAN(R)
This studyIntegrated DNA Technologies, Inc.TCTGGCTCTGGAGAAAGCTTATAACAGAGGATAACCGCGCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
Δcrp::KAN(R)
This studyIntegrated DNA Technologies, Inc.AAAATGGCGCGCTACCAGGTAACGCGCCACTCCGACGGGATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔcyaA::KAN(R)
This studyIntegrated DNA Technologies, Inc.GAATCACAGTCATGACGGGTAGCAAATCAGGCGATACGTCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔcyaA::KAN(R)
This studyIntegrated DNA Technologies, Inc.AGATTGCATGCCGGATAAGCCTCGCTTTCCGGCACGTTCATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔsucA::KAN(R)
This studyIntegrated DNA Technologies, Inc.ACGGCGAAGTAAGCATAAAAAAGATGCTTAAGGGATCACGGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔsucA::KAN(R)
This studyIntegrated DNA Technologies, Inc.GGTCAGGGACCAGAATATCTACGCTACTCATTGTGTAT
CCTTTATTTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
Δlpd::KAN(R)
This studyIntegrated DNA Technologies, Inc.GACGGGTATGACCGCC
GGAGATAAATATATAGAGGTCATGGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
Δlpd::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCCGCTTTTTTAATTGCCGGATGTTCCGGCAAACGAAAAATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔsucC::KAN(R)
This studyIntegrated DNA Technologies, Inc.GGTTTAAAAGATAACGATTACTGAAGGATGGACAGAACACGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔsucC::KAN(R)
This studyIntegrated DNA Technologies, Inc.TGGCAGATAACCTTGGTG
TTTTTATCGATTAAAATGGACATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔsdhA::KAN(R)
This studyIntegrated DNA Technologies, Inc.TTTACGTGATTTATG
GATTCGTTGTGGTGT
GGGGTGTGTGGTGTA
GGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔsdhA::KAN(R)
This studyIntegrated DNA Technologies, Inc.GATAAATTGAAAACT
CGAGTCTCATTTTCC
TGTCTCCGCATTAAC
GGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔgltA::KAN(R)
This studyIntegrated DNA Technologies, Inc.TAAGTTCCGGCAGTCTTACGCAATAAGGCGCTAAG
GAGACCTTAAGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔgltA::KAN(R)
This studyIntegrated DNA Technologies, Inc.CCCGCCATATGAACGGCGGGTTAAAATATTTACAACTTAGCAATCAACCATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔaceE::KAN(R)
This studyIntegrated DNA Technologies, Inc.GGTTCCAGAAAACTCAACGTTATTAGATAGATAAGGAATAACCCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔaceE::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCCCCGATGTCCGGTACTTTGATTTCGATAGCCATTATTCTTTTACCTCTTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
Δmdh::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCGGAGCAACATATCTTAGTTTATCAATATAATAAGGAGTTTAGGGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
Δmdh::KAN(R)
This studyIntegrated DNA Technologies, Inc.CCGGAGTCTGTGCTCCGGTTTTTTATTATCCGCTAATCAATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔnuoI::KAN(R)
This studyIntegrated DNA Technologies, Inc.CTGTCATTCTCTGGCAGGCGCAATAAGGGGCAATAAGACCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔnuoI::KAN(R)
This studyIntegrated DNA Technologies, Inc.AGGCCACAGATATAAAAAGC
GAACTCCATTGCCCCTCTCCTT
AACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔnuoM::KAN(R)
This studyIntegrated DNA Technologies, Inc.TCCGGTCCTGACGGGACTTTTACAAGGAATAAAGATCGCCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔnuoM::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCAGTGCGATCAGGTTTTGTGGAGTTATTGTCATGGCGATTTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔatpA::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCGCCTTGCAGACGTCTTGCAGTCTTAAGGGGACTGGAGCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔatpA::KAN(R)
This studyIntegrated DNA Technologies, Inc.TCAATGCCTTGCGGCCTGCCCTAAGGCAAGCCGCCAGACGTTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔatpB::KAN(R)
This studyIntegrated DNA Technologies, Inc.TGGCACCGGCTGTAATTAA
CAACAAAGGGTAAAAGGCATCGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔatpB::KAN(R)
This studyIntegrated DNA Technologies, Inc.CTCCAGTTTGTTTCAGTTAAAACGTAGTAGTGTTGGTAAATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔatpC::KAN(R)
This studyIntegrated DNA Technologies, Inc.GGAAAAAGCCAAAAAACTTTAACGCCTTAATCGGAGGGTGATGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔatpC::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCCTGTTTCCAGACTGGCTTTTGTGCTTTTCAAGCCGGTGTTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔatpD::KAN(R)
This studyIntegrated DNA Technologies, Inc.CCGCCGCGGTTTAAACAGGTT
ATTTCGTAGAGGATTTAAGGTGTA
GGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔatpD::KAN(R)
This studyIntegrated DNA Technologies, Inc.AGGTGGTAAGTCATTGCCATAT
CACCCTCCGATTAAGGCGTTAAC
GGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔfrdC::KAN(R)
This studyIntegrated DNA Technologies, Inc.TTTCTTATCGCGACCCTGAAACCACGCTAAGGAGTGCAACGTG
TAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔfrdC::KAN(R)
This studyIntegrated DNA Technologies, Inc.GTCAGAACGCTTTGGATTTGG
ATTAATCATCTCAGGCTCCTTAAC
GGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
Δpgi::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCTACAATCTTCCAAAGTCACA
ATTCTCAAAATCAGAAGAGTATTGC
TAGTGTAGGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
Δpgi::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCGGCGTGAACGCCTTATCC
GGCCTACATATCGACGATGATTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
Δzwf::KAN(R)
This studyIntegrated DNA Technologies, Inc.CTGGCTTAAGTACCGGGTTAG
TTAACTTAAGGAGAATGACGTGTA
GGCTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
Δzwf::KAN(R)
This studyIntegrated DNA Technologies, Inc.GCGCAAGATCATGTTACC
GGTAAAATAACCATAAAGGA
TAAGCGCAGATATTAACGGC
TGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔtktB::KAN(R)
This studyIntegrated DNA Technologies, Inc.CTTCTTGCCGCCAAACT
ATAAACCAGCCACGGAGTG
TTATGTGTAGGCTGGAGCTG
CTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔtktB::KAN(R)
This studyIntegrated DNA Technologies, Inc.GTCAGCGTCGCATCCGGCAA
TCAGCATCCGGCAATCACCATTA
ACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’)
ΔtalA::KAN(R)
This studyIntegrated DNA Technologies, Inc.
CGCACTCATCTAACACTTTACT
TTTCAAGGAGTATTTCCTGTGTAGG
CTGGAGCTGCTTC
Sequence-based reagentReverse Primer (5’ to 3’)
ΔtalA::KAN(R)
This studyIntegrated DNA Technologies, Inc.GGCAAGGTCTTTTCGGGAC
ATATAACACTCCGTGGCTGGT
TTAACGGCTGACATGGGAAT
Sequence-based reagentForward Primer (5’ to 3’) pUA66-crpThis studyIntegrated DNA Technologies, Inc.GCGCTCAGATCTTGATC
CGAAAGCTATGCTAAAACAGT
Sequence-based reagentReverse Primer (5’ to 3’) pUA66-crpThis studyIntegrated DNA Technologies, Inc.GCGCTCAGTACTttaAC
GAGTGCCGTAAACGA

Additional files

Supplementary file 1

MIC of antibiotics and concentrations of bactericidal antibiotics used in persister assays.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp1-v1.docx
Supplementary file 2

Analyzed metabolomics data.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp2-v1.docx
Supplementary file 3

Analyzed proteomics data.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp3-v1.docx
Supplementary file 4

Analyzed persister survival fraction data.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp4-v1.docx
Supplementary file 5

The knockout strains generated using the E. coli K-12 MG1655 background in this study.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp5-v1.docx
Supplementary file 6

Persister levels of E. coli K-12 MG1655 WT, Δcrp, and ΔcyaA strains in late stationary phase.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp6-v1.docx
Supplementary file 7

Persister levels of E. coli K-12 MG1655 (A) and BW25113 (B) WT, Δcrp, and ΔcyaA strains in the exponential growth phase.

https://cdn.elifesciences.org/articles/99735/elife-99735-supp7-v1.docx
MDAR checklist
https://cdn.elifesciences.org/articles/99735/elife-99735-mdarchecklist1-v1.docx

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  1. Han G Ngo
  2. Sayed Golam Mohiuddin
  3. Aina Ananda
  4. Mehmet Orman
(2025)
Unraveling CRP/cAMP-mediated metabolic regulation in Escherichia coli persister cells
eLife 13:RP99735.
https://doi.org/10.7554/eLife.99735.3