monarch pcr purification kit  (New England Biolabs)


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    Name:
    Monarch Genomic DNA Purification Kit
    Description:
    The Monarch Genomic DNA Purification Kit is a comprehensive solution for cell lysis RNA removal and purification of intact genomic DNA gDNA from a wide variety of biological samples including cultured cells blood and mammalian tissues Additionally bacteria and yeast can be processed with extra steps to enhance lysis in these tough to lyse samples Protocols are also included to enable purification from clinically relevant samples such as saliva and cheek swabs as well as rapid cleanup of previously extracted gDNA Purified gDNA has high quality metrics including A260 A280 1 8 and A260 A230 2 0 high DIN scores and minimal residual RNA The purified gDNA is suitable for downstream applications such as end point PCR qPCR and library prep for NGS It typically has a peak size of 50kb making this kit an excellent choice upstream of long read sequencing platforms
    Catalog Number:
    t3010l
    Price:
    395
    Size:
    150 preps
    Category:
    Genomic DNA Purification Kits
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    Structured Review

    New England Biolabs monarch pcr purification kit
    Monarch Genomic DNA Purification Kit
    The Monarch Genomic DNA Purification Kit is a comprehensive solution for cell lysis RNA removal and purification of intact genomic DNA gDNA from a wide variety of biological samples including cultured cells blood and mammalian tissues Additionally bacteria and yeast can be processed with extra steps to enhance lysis in these tough to lyse samples Protocols are also included to enable purification from clinically relevant samples such as saliva and cheek swabs as well as rapid cleanup of previously extracted gDNA Purified gDNA has high quality metrics including A260 A280 1 8 and A260 A230 2 0 high DIN scores and minimal residual RNA The purified gDNA is suitable for downstream applications such as end point PCR qPCR and library prep for NGS It typically has a peak size of 50kb making this kit an excellent choice upstream of long read sequencing platforms
    https://www.bioz.com/result/monarch pcr purification kit/product/New England Biolabs
    Average 97 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    monarch pcr purification kit - by Bioz Stars, 2020-09
    97/100 stars

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    Images

    1) Product Images from "Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics"

    Article Title: Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics

    Journal: bioRxiv

    doi: 10.1101/2020.04.20.049437

    PYO induces expression of specific efflux systems, conferring cross-tolerance to fluoroquinolones. A . Structures of PYO, two representative fluoroquinolones (CP = ciprofloxacin, LV = levofloxacin) and two representative aminoglycosides (GM = gentamicin, TM = tobramycin). PYO and fluoroquinolones are pumped by MexEF-OprN and MexGHI-OpmD, while aminoglycosides are not 11 . Rings with an aromatic character are highlighted in red. B . Normalized cDNA levels for genes within operons coding for the 11 main RND efflux systems in P. aeruginosa (left). PYO-dose-dependent changes in expression of mexEF-oprN and mexGHI- opmD systems (right; n = 3). For full qRT-PCR dataset, see Figs. S2, S3 and S4. C . Effect of PYO on tolerance to CP and LV in glucose minimal medium (left), and to CP in SCFM (right) (all 1 µg/mL) (n = 4). PYO itself was not toxic under the experimental conditions 8 . WT made 50-80 µM PYO as measured by absorbance of the culture supernatant at 691 nm. See Fig. S5A for experimental design. D-E . Effect of PYO on lag during outgrowth after exposure to CP. A representative field of view over different time points (D; magenta = WT::mApple, green = Δ phz ::GFP; see Movie S1) is shown together with the quantification of growth area on the agarose pads at time 0 hrs and 15 hrs (E). For these experiments, a culture of each strain tested was grown and exposed to CP (10 µg/mL) separately, then cells of both cultures were washed, mixed and placed together on a pad and imaged during outgrowth. The pads did not contain any PYO or CP (see Methods and Fig. S5D for details). White arrows in the displayed images point to regions with faster recovery of WT growth. The field of view displayed is marked with a black arrow in the quantification plot. The results for the experiment with swapped fluorescent proteins are shown in Fig. S5E. Scale bar: 20 µm. F . Tolerance of Δ phz to CP (1 µg/mL) in stationary phase in the presence of different concentrations of PYO (n = 4). G . Tolerance of Δ phz to CP (1 µg/mL) upon artificial induction of the mexGHI-opmD operon with arabinose (n = 4). The dashed green line marks the average survival of PYO-producing WT under similar conditions (without arabinose). Statistics: C, F – One-way ANOVA with Tukey’s HSD multiple-comparison test, with asterisks showing significant differences relative to untreated Δ phz (no PYO); E, G – Welch’s unpaired t- test (* p
    Figure Legend Snippet: PYO induces expression of specific efflux systems, conferring cross-tolerance to fluoroquinolones. A . Structures of PYO, two representative fluoroquinolones (CP = ciprofloxacin, LV = levofloxacin) and two representative aminoglycosides (GM = gentamicin, TM = tobramycin). PYO and fluoroquinolones are pumped by MexEF-OprN and MexGHI-OpmD, while aminoglycosides are not 11 . Rings with an aromatic character are highlighted in red. B . Normalized cDNA levels for genes within operons coding for the 11 main RND efflux systems in P. aeruginosa (left). PYO-dose-dependent changes in expression of mexEF-oprN and mexGHI- opmD systems (right; n = 3). For full qRT-PCR dataset, see Figs. S2, S3 and S4. C . Effect of PYO on tolerance to CP and LV in glucose minimal medium (left), and to CP in SCFM (right) (all 1 µg/mL) (n = 4). PYO itself was not toxic under the experimental conditions 8 . WT made 50-80 µM PYO as measured by absorbance of the culture supernatant at 691 nm. See Fig. S5A for experimental design. D-E . Effect of PYO on lag during outgrowth after exposure to CP. A representative field of view over different time points (D; magenta = WT::mApple, green = Δ phz ::GFP; see Movie S1) is shown together with the quantification of growth area on the agarose pads at time 0 hrs and 15 hrs (E). For these experiments, a culture of each strain tested was grown and exposed to CP (10 µg/mL) separately, then cells of both cultures were washed, mixed and placed together on a pad and imaged during outgrowth. The pads did not contain any PYO or CP (see Methods and Fig. S5D for details). White arrows in the displayed images point to regions with faster recovery of WT growth. The field of view displayed is marked with a black arrow in the quantification plot. The results for the experiment with swapped fluorescent proteins are shown in Fig. S5E. Scale bar: 20 µm. F . Tolerance of Δ phz to CP (1 µg/mL) in stationary phase in the presence of different concentrations of PYO (n = 4). G . Tolerance of Δ phz to CP (1 µg/mL) upon artificial induction of the mexGHI-opmD operon with arabinose (n = 4). The dashed green line marks the average survival of PYO-producing WT under similar conditions (without arabinose). Statistics: C, F – One-way ANOVA with Tukey’s HSD multiple-comparison test, with asterisks showing significant differences relative to untreated Δ phz (no PYO); E, G – Welch’s unpaired t- test (* p

    Techniques Used: Expressing, Quantitative RT-PCR

    Related Articles

    DNA Extraction:

    Article Title: Complete Genome Sequence of Bacillus velezensis Strain S4, Isolated from Biochar-Treated Soil
    Article Snippet: .. DNA was extracted from 3.5 ml of this culture (Monarch DNA extraction kit, NEB product number T3010). .. Single-molecule real-time (SMRT) libraries were prepared using the standard PacBio protocol for 20-kb libraries.

    Article Title: lncRNA MEG8 Upregulates miR-770-5p Through Methylation and Promotes Cell Apoptosis in Diabetic Nephropathy
    Article Snippet: .. Methylation-Specific PCR (MSP) CIHP-1 cells were subjected to genomic DNA isolation using Monarch® Genomic DNA Purification Kit (NEB). .. EZ DNA Methylation-GoldTM kit (ZYMO) was used to convert DNA samples.

    Nucleic Acid Electrophoresis:

    Article Title: Rates and Microbial Players of Iron-Driven Anaerobic Oxidation of Methane in Methanic Marine Sediments
    Article Snippet: .. PCR cycling conditions include: 95°C: 5 min; 28 cycles at 98°C: 20 s, 60°C: 20 s, 72°C: 20 s; 72°C: 1 min. PCR products were screened by gel electrophoresis as mentioned before and purified using Monarch® PCR & DNA purification kit (New England Biolabs, Germany). .. PCR products were quantified using Quant-iT PicoGreen dsDNA assay kit (Invitrogen-Thermo Fischer Scientific, Steinheim, Germany).

    Amplification:

    Article Title: Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics
    Article Snippet: .. Fragments amplified from P. aeruginosa PA14 genomic DNA (gDNA) and cleaned up using the Monarch PCR Purification kit (New England Biolabs) were used for Gibson assembly together with pMQ30 cut with SacI and HindIII. .. The assembled construct was then transformed into Escherichia coli DH10B, with transformants being selected in LB with 20 µg/mL gentamicin.

    Article Title: A High-efficacy CRISPRi System for Gene Function Discovery in Zymomonas mobilis
    Article Snippet: .. For method two, fragments were amplified by PCR with primers oJMP197 and oJMP198 from a 78 nt oligonucleotide followed by digestion with BsaI-HF-v2 and purification with the Monarch DNA purification kit (NEB) following the manufacturer’s oligonucleotide purification protocol. .. Inserts (2 μl of a 1:40 dilution of annealed oligos or 2 ng purified digested PCR product) were ligated into 50 ng BsaI-digested vector.

    Methylation:

    Article Title: lncRNA MEG8 Upregulates miR-770-5p Through Methylation and Promotes Cell Apoptosis in Diabetic Nephropathy
    Article Snippet: .. Methylation-Specific PCR (MSP) CIHP-1 cells were subjected to genomic DNA isolation using Monarch® Genomic DNA Purification Kit (NEB). .. EZ DNA Methylation-GoldTM kit (ZYMO) was used to convert DNA samples.

    Isolation:

    Article Title: An efficient CRISPR-based strategy to insert small and large fragments of DNA using short homology arms
    Article Snippet: .. Reaction products were pooled in two samples and ssDNA isolated using NEB Monarch DNA purification kit following the manufacturer’s instructions (NEB #T1030L). .. Samples were isolated in 2 × 10 μl prewarmed (55°C) elution buffer from the kit and the DNA concentration was measured using NanoDrop One (ThermoFisher Scientific).

    Purification:

    Article Title: Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics
    Article Snippet: .. Fragments amplified from P. aeruginosa PA14 genomic DNA (gDNA) and cleaned up using the Monarch PCR Purification kit (New England Biolabs) were used for Gibson assembly together with pMQ30 cut with SacI and HindIII. .. The assembled construct was then transformed into Escherichia coli DH10B, with transformants being selected in LB with 20 µg/mL gentamicin.

    Article Title: A High-efficacy CRISPRi System for Gene Function Discovery in Zymomonas mobilis
    Article Snippet: .. For method two, fragments were amplified by PCR with primers oJMP197 and oJMP198 from a 78 nt oligonucleotide followed by digestion with BsaI-HF-v2 and purification with the Monarch DNA purification kit (NEB) following the manufacturer’s oligonucleotide purification protocol. .. Inserts (2 μl of a 1:40 dilution of annealed oligos or 2 ng purified digested PCR product) were ligated into 50 ng BsaI-digested vector.

    Article Title: Potent CRISPR-Cas9 inhibitors from Staphylococcus genomes
    Article Snippet: .. Cells were then treated with lysostaphin for 1 h at 37 °C. gDNA was then extracted using the Monarch gDNA Purification Kit from NEB following the manufacturer’s instructions. ..

    Article Title: Macrovesicular steatosis in nonalcoholic fatty liver disease is a consequence of purine nucleotide cycle driven fumarate accumulation
    Article Snippet: .. DNA hydroxymethylation immunoprecipitation and sequencing (hmeDIP-sequencing) DNA was purified using the Monarch® Genomic DNA Purification kit (New England BioLabs, T3010S). .. DNA immunoprecipitation and sequencing was performed as previously described, using the Ion Proton platform , with the addition of an IgG control (Merck, 12-370).

    Article Title: Rates and Microbial Players of Iron-Driven Anaerobic Oxidation of Methane in Methanic Marine Sediments
    Article Snippet: .. PCR cycling conditions include: 95°C: 5 min; 28 cycles at 98°C: 20 s, 60°C: 20 s, 72°C: 20 s; 72°C: 1 min. PCR products were screened by gel electrophoresis as mentioned before and purified using Monarch® PCR & DNA purification kit (New England Biolabs, Germany). .. PCR products were quantified using Quant-iT PicoGreen dsDNA assay kit (Invitrogen-Thermo Fischer Scientific, Steinheim, Germany).

    Immunoprecipitation:

    Article Title: Macrovesicular steatosis in nonalcoholic fatty liver disease is a consequence of purine nucleotide cycle driven fumarate accumulation
    Article Snippet: .. DNA hydroxymethylation immunoprecipitation and sequencing (hmeDIP-sequencing) DNA was purified using the Monarch® Genomic DNA Purification kit (New England BioLabs, T3010S). .. DNA immunoprecipitation and sequencing was performed as previously described, using the Ion Proton platform , with the addition of an IgG control (Merck, 12-370).

    Polymerase Chain Reaction:

    Article Title: Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics
    Article Snippet: .. Fragments amplified from P. aeruginosa PA14 genomic DNA (gDNA) and cleaned up using the Monarch PCR Purification kit (New England Biolabs) were used for Gibson assembly together with pMQ30 cut with SacI and HindIII. .. The assembled construct was then transformed into Escherichia coli DH10B, with transformants being selected in LB with 20 µg/mL gentamicin.

    Article Title: A High-efficacy CRISPRi System for Gene Function Discovery in Zymomonas mobilis
    Article Snippet: .. For method two, fragments were amplified by PCR with primers oJMP197 and oJMP198 from a 78 nt oligonucleotide followed by digestion with BsaI-HF-v2 and purification with the Monarch DNA purification kit (NEB) following the manufacturer’s oligonucleotide purification protocol. .. Inserts (2 μl of a 1:40 dilution of annealed oligos or 2 ng purified digested PCR product) were ligated into 50 ng BsaI-digested vector.

    Article Title: Rates and Microbial Players of Iron-Driven Anaerobic Oxidation of Methane in Methanic Marine Sediments
    Article Snippet: .. PCR cycling conditions include: 95°C: 5 min; 28 cycles at 98°C: 20 s, 60°C: 20 s, 72°C: 20 s; 72°C: 1 min. PCR products were screened by gel electrophoresis as mentioned before and purified using Monarch® PCR & DNA purification kit (New England Biolabs, Germany). .. PCR products were quantified using Quant-iT PicoGreen dsDNA assay kit (Invitrogen-Thermo Fischer Scientific, Steinheim, Germany).

    Article Title: lncRNA MEG8 Upregulates miR-770-5p Through Methylation and Promotes Cell Apoptosis in Diabetic Nephropathy
    Article Snippet: .. Methylation-Specific PCR (MSP) CIHP-1 cells were subjected to genomic DNA isolation using Monarch® Genomic DNA Purification Kit (NEB). .. EZ DNA Methylation-GoldTM kit (ZYMO) was used to convert DNA samples.

    DNA Purification:

    Article Title: A High-efficacy CRISPRi System for Gene Function Discovery in Zymomonas mobilis
    Article Snippet: .. For method two, fragments were amplified by PCR with primers oJMP197 and oJMP198 from a 78 nt oligonucleotide followed by digestion with BsaI-HF-v2 and purification with the Monarch DNA purification kit (NEB) following the manufacturer’s oligonucleotide purification protocol. .. Inserts (2 μl of a 1:40 dilution of annealed oligos or 2 ng purified digested PCR product) were ligated into 50 ng BsaI-digested vector.

    Article Title: Macrovesicular steatosis in nonalcoholic fatty liver disease is a consequence of purine nucleotide cycle driven fumarate accumulation
    Article Snippet: .. DNA hydroxymethylation immunoprecipitation and sequencing (hmeDIP-sequencing) DNA was purified using the Monarch® Genomic DNA Purification kit (New England BioLabs, T3010S). .. DNA immunoprecipitation and sequencing was performed as previously described, using the Ion Proton platform , with the addition of an IgG control (Merck, 12-370).

    Article Title: An efficient CRISPR-based strategy to insert small and large fragments of DNA using short homology arms
    Article Snippet: .. Reaction products were pooled in two samples and ssDNA isolated using NEB Monarch DNA purification kit following the manufacturer’s instructions (NEB #T1030L). .. Samples were isolated in 2 × 10 μl prewarmed (55°C) elution buffer from the kit and the DNA concentration was measured using NanoDrop One (ThermoFisher Scientific).

    Article Title: Rates and Microbial Players of Iron-Driven Anaerobic Oxidation of Methane in Methanic Marine Sediments
    Article Snippet: .. PCR cycling conditions include: 95°C: 5 min; 28 cycles at 98°C: 20 s, 60°C: 20 s, 72°C: 20 s; 72°C: 1 min. PCR products were screened by gel electrophoresis as mentioned before and purified using Monarch® PCR & DNA purification kit (New England Biolabs, Germany). .. PCR products were quantified using Quant-iT PicoGreen dsDNA assay kit (Invitrogen-Thermo Fischer Scientific, Steinheim, Germany).

    Article Title: lncRNA MEG8 Upregulates miR-770-5p Through Methylation and Promotes Cell Apoptosis in Diabetic Nephropathy
    Article Snippet: .. Methylation-Specific PCR (MSP) CIHP-1 cells were subjected to genomic DNA isolation using Monarch® Genomic DNA Purification Kit (NEB). .. EZ DNA Methylation-GoldTM kit (ZYMO) was used to convert DNA samples.

    Sequencing:

    Article Title: Macrovesicular steatosis in nonalcoholic fatty liver disease is a consequence of purine nucleotide cycle driven fumarate accumulation
    Article Snippet: .. DNA hydroxymethylation immunoprecipitation and sequencing (hmeDIP-sequencing) DNA was purified using the Monarch® Genomic DNA Purification kit (New England BioLabs, T3010S). .. DNA immunoprecipitation and sequencing was performed as previously described, using the Ion Proton platform , with the addition of an IgG control (Merck, 12-370).

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    New England Biolabs monarch pcr purification kit
    PYO induces expression of specific efflux systems, conferring cross-tolerance to fluoroquinolones. A . Structures of PYO, two representative fluoroquinolones (CP = ciprofloxacin, LV = levofloxacin) and two representative aminoglycosides (GM = gentamicin, TM = tobramycin). PYO and fluoroquinolones are pumped by MexEF-OprN and MexGHI-OpmD, while aminoglycosides are not 11 . Rings with an aromatic character are highlighted in red. B . Normalized cDNA levels for genes within operons coding for the 11 main RND efflux systems in P. <t>aeruginosa</t> (left). PYO-dose-dependent changes in expression of mexEF-oprN and mexGHI- opmD systems (right; n = 3). For full <t>qRT-PCR</t> dataset, see Figs. S2, S3 and S4. C . Effect of PYO on tolerance to CP and LV in glucose minimal medium (left), and to CP in SCFM (right) (all 1 µg/mL) (n = 4). PYO itself was not toxic under the experimental conditions 8 . WT made 50-80 µM PYO as measured by absorbance of the culture supernatant at 691 nm. See Fig. S5A for experimental design. D-E . Effect of PYO on lag during outgrowth after exposure to CP. A representative field of view over different time points (D; magenta = WT::mApple, green = Δ phz ::GFP; see Movie S1) is shown together with the quantification of growth area on the agarose pads at time 0 hrs and 15 hrs (E). For these experiments, a culture of each strain tested was grown and exposed to CP (10 µg/mL) separately, then cells of both cultures were washed, mixed and placed together on a pad and imaged during outgrowth. The pads did not contain any PYO or CP (see Methods and Fig. S5D for details). White arrows in the displayed images point to regions with faster recovery of WT growth. The field of view displayed is marked with a black arrow in the quantification plot. The results for the experiment with swapped fluorescent proteins are shown in Fig. S5E. Scale bar: 20 µm. F . Tolerance of Δ phz to CP (1 µg/mL) in stationary phase in the presence of different concentrations of PYO (n = 4). G . Tolerance of Δ phz to CP (1 µg/mL) upon artificial induction of the mexGHI-opmD operon with arabinose (n = 4). The dashed green line marks the average survival of PYO-producing WT under similar conditions (without arabinose). Statistics: C, F – One-way ANOVA with Tukey’s HSD multiple-comparison test, with asterisks showing significant differences relative to untreated Δ phz (no PYO); E, G – Welch’s unpaired t- test (* p
    Monarch Pcr Purification Kit, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/monarch pcr purification kit/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    monarch pcr purification kit - by Bioz Stars, 2020-09
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    PYO induces expression of specific efflux systems, conferring cross-tolerance to fluoroquinolones. A . Structures of PYO, two representative fluoroquinolones (CP = ciprofloxacin, LV = levofloxacin) and two representative aminoglycosides (GM = gentamicin, TM = tobramycin). PYO and fluoroquinolones are pumped by MexEF-OprN and MexGHI-OpmD, while aminoglycosides are not 11 . Rings with an aromatic character are highlighted in red. B . Normalized cDNA levels for genes within operons coding for the 11 main RND efflux systems in P. aeruginosa (left). PYO-dose-dependent changes in expression of mexEF-oprN and mexGHI- opmD systems (right; n = 3). For full qRT-PCR dataset, see Figs. S2, S3 and S4. C . Effect of PYO on tolerance to CP and LV in glucose minimal medium (left), and to CP in SCFM (right) (all 1 µg/mL) (n = 4). PYO itself was not toxic under the experimental conditions 8 . WT made 50-80 µM PYO as measured by absorbance of the culture supernatant at 691 nm. See Fig. S5A for experimental design. D-E . Effect of PYO on lag during outgrowth after exposure to CP. A representative field of view over different time points (D; magenta = WT::mApple, green = Δ phz ::GFP; see Movie S1) is shown together with the quantification of growth area on the agarose pads at time 0 hrs and 15 hrs (E). For these experiments, a culture of each strain tested was grown and exposed to CP (10 µg/mL) separately, then cells of both cultures were washed, mixed and placed together on a pad and imaged during outgrowth. The pads did not contain any PYO or CP (see Methods and Fig. S5D for details). White arrows in the displayed images point to regions with faster recovery of WT growth. The field of view displayed is marked with a black arrow in the quantification plot. The results for the experiment with swapped fluorescent proteins are shown in Fig. S5E. Scale bar: 20 µm. F . Tolerance of Δ phz to CP (1 µg/mL) in stationary phase in the presence of different concentrations of PYO (n = 4). G . Tolerance of Δ phz to CP (1 µg/mL) upon artificial induction of the mexGHI-opmD operon with arabinose (n = 4). The dashed green line marks the average survival of PYO-producing WT under similar conditions (without arabinose). Statistics: C, F – One-way ANOVA with Tukey’s HSD multiple-comparison test, with asterisks showing significant differences relative to untreated Δ phz (no PYO); E, G – Welch’s unpaired t- test (* p

    Journal: bioRxiv

    Article Title: Bacterial defenses against a natural antibiotic promote collateral resilience to clinical antibiotics

    doi: 10.1101/2020.04.20.049437

    Figure Lengend Snippet: PYO induces expression of specific efflux systems, conferring cross-tolerance to fluoroquinolones. A . Structures of PYO, two representative fluoroquinolones (CP = ciprofloxacin, LV = levofloxacin) and two representative aminoglycosides (GM = gentamicin, TM = tobramycin). PYO and fluoroquinolones are pumped by MexEF-OprN and MexGHI-OpmD, while aminoglycosides are not 11 . Rings with an aromatic character are highlighted in red. B . Normalized cDNA levels for genes within operons coding for the 11 main RND efflux systems in P. aeruginosa (left). PYO-dose-dependent changes in expression of mexEF-oprN and mexGHI- opmD systems (right; n = 3). For full qRT-PCR dataset, see Figs. S2, S3 and S4. C . Effect of PYO on tolerance to CP and LV in glucose minimal medium (left), and to CP in SCFM (right) (all 1 µg/mL) (n = 4). PYO itself was not toxic under the experimental conditions 8 . WT made 50-80 µM PYO as measured by absorbance of the culture supernatant at 691 nm. See Fig. S5A for experimental design. D-E . Effect of PYO on lag during outgrowth after exposure to CP. A representative field of view over different time points (D; magenta = WT::mApple, green = Δ phz ::GFP; see Movie S1) is shown together with the quantification of growth area on the agarose pads at time 0 hrs and 15 hrs (E). For these experiments, a culture of each strain tested was grown and exposed to CP (10 µg/mL) separately, then cells of both cultures were washed, mixed and placed together on a pad and imaged during outgrowth. The pads did not contain any PYO or CP (see Methods and Fig. S5D for details). White arrows in the displayed images point to regions with faster recovery of WT growth. The field of view displayed is marked with a black arrow in the quantification plot. The results for the experiment with swapped fluorescent proteins are shown in Fig. S5E. Scale bar: 20 µm. F . Tolerance of Δ phz to CP (1 µg/mL) in stationary phase in the presence of different concentrations of PYO (n = 4). G . Tolerance of Δ phz to CP (1 µg/mL) upon artificial induction of the mexGHI-opmD operon with arabinose (n = 4). The dashed green line marks the average survival of PYO-producing WT under similar conditions (without arabinose). Statistics: C, F – One-way ANOVA with Tukey’s HSD multiple-comparison test, with asterisks showing significant differences relative to untreated Δ phz (no PYO); E, G – Welch’s unpaired t- test (* p

    Article Snippet: Fragments amplified from P. aeruginosa PA14 genomic DNA (gDNA) and cleaned up using the Monarch PCR Purification kit (New England Biolabs) were used for Gibson assembly together with pMQ30 cut with SacI and HindIII.

    Techniques: Expressing, Quantitative RT-PCR