escherichia coli dh5α  (Thermo Fisher)


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    Structured Review

    Thermo Fisher escherichia coli dh5α
    ( a ) Remote effect of emissions of the PGPB Azospirillum brasilense Cd and Bacillus pumilus ES4 and the control bacterium <t>Escherichia</t> coli <t>DH5α</t> on growth of the microalgae Chlorella sorokiniana , ( b ) growth in the presence of the CO 2 absorbant LiOH•H 2 O, 0.3 g and ( c ) 0.5 g. ( d ) Remote effect of emissions on the volume of C. sorokiniana cells, ( e ) Growth of the PGPB in the culture medium, and ( f ), Increase in pH in the medium. The control bacterium ( E. coli ) was used only in experiments measuring the potential effect of produced CO 2 on microalgae growth and metabolite production. Values on curves denoted by different capital letters differ significantly using one-way ANOVA combined with LSD post-hoc analysis at P
    Escherichia Coli Dh5α, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 154 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/escherichia coli dh5α/product/Thermo Fisher
    Average 94 stars, based on 154 article reviews
    Price from $9.99 to $1999.99
    escherichia coli dh5α - by Bioz Stars, 2020-07
    94/100 stars

    Images

    1) Product Images from "Enhanced performance of the microalga Chlorella sorokiniana remotely induced by the plant growth-promoting bacteria Azospirillum brasilense and Bacillus pumilus"

    Article Title: Enhanced performance of the microalga Chlorella sorokiniana remotely induced by the plant growth-promoting bacteria Azospirillum brasilense and Bacillus pumilus

    Journal: Scientific Reports

    doi: 10.1038/srep41310

    ( a ) Remote effect of emissions of the PGPB Azospirillum brasilense Cd and Bacillus pumilus ES4 and the control bacterium Escherichia coli DH5α on growth of the microalgae Chlorella sorokiniana , ( b ) growth in the presence of the CO 2 absorbant LiOH•H 2 O, 0.3 g and ( c ) 0.5 g. ( d ) Remote effect of emissions on the volume of C. sorokiniana cells, ( e ) Growth of the PGPB in the culture medium, and ( f ), Increase in pH in the medium. The control bacterium ( E. coli ) was used only in experiments measuring the potential effect of produced CO 2 on microalgae growth and metabolite production. Values on curves denoted by different capital letters differ significantly using one-way ANOVA combined with LSD post-hoc analysis at P
    Figure Legend Snippet: ( a ) Remote effect of emissions of the PGPB Azospirillum brasilense Cd and Bacillus pumilus ES4 and the control bacterium Escherichia coli DH5α on growth of the microalgae Chlorella sorokiniana , ( b ) growth in the presence of the CO 2 absorbant LiOH•H 2 O, 0.3 g and ( c ) 0.5 g. ( d ) Remote effect of emissions on the volume of C. sorokiniana cells, ( e ) Growth of the PGPB in the culture medium, and ( f ), Increase in pH in the medium. The control bacterium ( E. coli ) was used only in experiments measuring the potential effect of produced CO 2 on microalgae growth and metabolite production. Values on curves denoted by different capital letters differ significantly using one-way ANOVA combined with LSD post-hoc analysis at P

    Techniques Used: Produced

    2) Product Images from "Non-contact positions impose site selectivity on Cre recombinase"

    Article Title: Non-contact positions impose site selectivity on Cre recombinase

    Journal: Nucleic Acids Research

    doi:

    Modulation of Cre-E262G recombination at mutant lox sites. Shown are recombination frequencies for wt and the indicated mutant Cre recombinases at loxP and at mutant lox sites that differ from loxP at positions 11 and 12. Escherichia coli DH5α carrying a resident pBAD construct expressing the indicated cre mutant was transformed with the indicated lox 2 neo plasmids and cre expression induced for 1 h before plating non-selectively to yield about 200 colonies/experiment. Sensitivity to kanamycin, signifying excisive recombination, was determined by replica plating. The frequencies shown are calculated from at least two independent experiments.
    Figure Legend Snippet: Modulation of Cre-E262G recombination at mutant lox sites. Shown are recombination frequencies for wt and the indicated mutant Cre recombinases at loxP and at mutant lox sites that differ from loxP at positions 11 and 12. Escherichia coli DH5α carrying a resident pBAD construct expressing the indicated cre mutant was transformed with the indicated lox 2 neo plasmids and cre expression induced for 1 h before plating non-selectively to yield about 200 colonies/experiment. Sensitivity to kanamycin, signifying excisive recombination, was determined by replica plating. The frequencies shown are calculated from at least two independent experiments.

    Techniques Used: Mutagenesis, Construct, Expressing, Transformation Assay

    Related Articles

    Negative Control:

    Article Title: Enhanced performance of the microalga Chlorella sorokiniana remotely induced by the plant growth-promoting bacteria Azospirillum brasilense and Bacillus pumilus
    Article Snippet: .. In experiments that measured the potential effect of CO2 , Escherichia coli DH5α (Invitrogen, Carlsbad, CA) served as the negative control for microalgal growth and promoting metabolites because it does not have any plant growth-promoting effects; it also served as a positive control in the CO2 experiment because it produces CO2 , as any E. coli . .. For initial culturing of the microalga, 10 mL axenic C. sorokiniana culture, cultivated in sterile mineral medium (C30), was added to a sterile flask containing 90 mL sterile C30 medium, composed of (in g·L−1 ): KNO3 (25), MgSO4 •7H2 O (10), KH2 PO4 (4), K2 HPO4 (1), FeSO4 •7H2 O (1), and (in μg·L−1 ): H3 BO3 (2.86), MnCl2 •4H2 O (1.81), ZnSO4 •7H2 O (0.11), CuSO4 •5H2 O (0.09), NaMoO4 (0.021), pH 5.25 and incubated at 27 ± 2 °C on a rotary shaker at 120 rpm under 60 μmol photon·m−2 ·s−1 continuous light intensity for 6 days .

    Positive Control:

    Article Title: Maturation of molybdoenzymes and its influence on the pathogenesis of non-typeable Haemophilus influenzae
    Article Snippet: .. E. coli Dh5α (Life Technologies) was used as a positive control. ..

    Article Title: Enhanced performance of the microalga Chlorella sorokiniana remotely induced by the plant growth-promoting bacteria Azospirillum brasilense and Bacillus pumilus
    Article Snippet: .. In experiments that measured the potential effect of CO2 , Escherichia coli DH5α (Invitrogen, Carlsbad, CA) served as the negative control for microalgal growth and promoting metabolites because it does not have any plant growth-promoting effects; it also served as a positive control in the CO2 experiment because it produces CO2 , as any E. coli . .. For initial culturing of the microalga, 10 mL axenic C. sorokiniana culture, cultivated in sterile mineral medium (C30), was added to a sterile flask containing 90 mL sterile C30 medium, composed of (in g·L−1 ): KNO3 (25), MgSO4 •7H2 O (10), KH2 PO4 (4), K2 HPO4 (1), FeSO4 •7H2 O (1), and (in μg·L−1 ): H3 BO3 (2.86), MnCl2 •4H2 O (1.81), ZnSO4 •7H2 O (0.11), CuSO4 •5H2 O (0.09), NaMoO4 (0.021), pH 5.25 and incubated at 27 ± 2 °C on a rotary shaker at 120 rpm under 60 μmol photon·m−2 ·s−1 continuous light intensity for 6 days .

    Ligation:

    Article Title: Functions of the Mismatch Repair Gene mutS from Acinetobacter sp. Strain ADP1 †
    Article Snippet: .. Recombinant plasmids were isolated by transforming E. coli DH5α with the appropriate ligation reaction according to the transformation protocol provided by the supplier (Gibco BRL). .. Plasmid pZR7000 was constructed by blunt end ligation of the MUTSF2-MUTSR3 PCR product into the Sma I site of pGEM-3Zf(+).

    Isolation:

    Article Title: Functions of the Mismatch Repair Gene mutS from Acinetobacter sp. Strain ADP1 †
    Article Snippet: .. Recombinant plasmids were isolated by transforming E. coli DH5α with the appropriate ligation reaction according to the transformation protocol provided by the supplier (Gibco BRL). .. Plasmid pZR7000 was constructed by blunt end ligation of the MUTSF2-MUTSR3 PCR product into the Sma I site of pGEM-3Zf(+).

    Construct:

    Article Title: Bacillus thuringiensis subsp. kurstaki HD1 as a factory to synthesize alkali-labile ChiA74?sp chitinase inclusions, Cry crystals and spores for applied use
    Article Snippet: .. All constructs (Figure A) were propagated in E. coli DH5α [supE44, DlacU169 (F80lacZDM15 ) hsdR17 recA1 end A1 gyrA96 thi-1 relA1 ] (Invitrogen, Carlsbad CA, USA) and then used for transforming the acrystalliferous strain of B. thuringiensis subsp. israelensis 4Q7, (hereafter 4Q7), and B. thuringiensis subsp. kurstaki HD1 (hereafter HD1) (Bacillus Genetic Stock Center, Columbus, OH). .. Plasmid pGLO is a vector that harbors the green fluorescent protein (gfp ) gene under the control arabinose (araC ) promoter and contains an ampicillin (bla ) resistance gene marker (Bio-Rad, Hercules CA, USA).

    Article Title: Improving Acetate Tolerance of Escherichia coli by Rewiring Its Global Regulator cAMP Receptor Protein (CRP)
    Article Snippet: .. Materials The host strain E. coli DH5α ∆crp was constructed by knocking out crp from E. coli DH5α (Invitrogen, San Diego, US) according to previously established protocol [ ]. .. Overnight culture was prepared in Luria-Bertani (LB) medium containing 1% tryptone (Oxoid, Hampshire, UK), 0.5% yeast extract (Merck, Damstadt, Germany), and 1% sodium chloride (Merck, Damstadt, Germany).

    Real-time Polymerase Chain Reaction:

    Article Title: Natural microbial communities supporting the transfer of the IncP-1β plasmid pB10 exhibit a higher initial content of plasmids from the same incompatibility group
    Article Snippet: .. Basically, sets of qPCR primers were designed to prime on both sides of a unique junction between building blocks of the element targeted for quantification: a unique assembly between two truncated transposons for plasmid pB10 (target coordinates 45968–46102; GenBank NC_004840), and both sides of a unique 97 kb-deletion in the chromosome of E. coli DH5α (target coordinates on K12 274654–372933; GenBank U00096; ; ). qPCRs were performed in triplicate using a “Step One Plus Real-Time PCR System” (Applied Biosystems, driver: StepOne Software v2.2) with thermocycling conditions set as follows: 2 min at 50°C, then 10 min at 95°C followed by 45 cycles of 15 s at 95°C and 1 min at 60°C. .. Quantifications were carried out from 25 ng of community DNA using a “TaqMan® Universal PCR Master Mix, NoAmpErase® UNG” (Applied Biosystems) as recommended by the manufacturer, with 800 nM of each primer, and 300 nM of TaqMan probe, in a 25 μL reaction volume.

    Expressing:

    Article Title: Amino acid residues 201-205 in C-terminal acidic tail region plays a crucial role in antibacterial activity of HMGB1
    Article Snippet: .. Briefly, the lysates of E. coli DH5α expressing the corresponding proteins were subjected to SDS-PAGE and then transferred onto polyvinylidene difluoride membranes (Pierce). .. After blocked with 5% non-fat milk in phosphate buffered saline containing 0.1% Tween 20 (PBS-T), the membranes were incubated with mouse anti-His monoclonal antibody (Qiagen).

    Transformation Assay:

    Article Title: Requirement of MrpH for Mannose-Resistant Proteus-Like Fimbria-Mediated Hemagglutination by Proteus mirabilis
    Article Snippet: .. The isopropyl-β- d -thiogalactopyranoside (IPTG)-induced cell lysate (about 10 mg protein) of E. coli DH5α transformed with plasmid pMAL-C2 was coupled to an AminoLink Plus column (Pierce Inc., Rockford, Ill.) as specified by the manufacturer. ..

    Article Title: Functions of the Mismatch Repair Gene mutS from Acinetobacter sp. Strain ADP1 †
    Article Snippet: .. Recombinant plasmids were isolated by transforming E. coli DH5α with the appropriate ligation reaction according to the transformation protocol provided by the supplier (Gibco BRL). .. Plasmid pZR7000 was constructed by blunt end ligation of the MUTSF2-MUTSR3 PCR product into the Sma I site of pGEM-3Zf(+).

    Recombinant:

    Article Title: Functions of the Mismatch Repair Gene mutS from Acinetobacter sp. Strain ADP1 †
    Article Snippet: .. Recombinant plasmids were isolated by transforming E. coli DH5α with the appropriate ligation reaction according to the transformation protocol provided by the supplier (Gibco BRL). .. Plasmid pZR7000 was constructed by blunt end ligation of the MUTSF2-MUTSR3 PCR product into the Sma I site of pGEM-3Zf(+).

    SDS Page:

    Article Title: Amino acid residues 201-205 in C-terminal acidic tail region plays a crucial role in antibacterial activity of HMGB1
    Article Snippet: .. Briefly, the lysates of E. coli DH5α expressing the corresponding proteins were subjected to SDS-PAGE and then transferred onto polyvinylidene difluoride membranes (Pierce). .. After blocked with 5% non-fat milk in phosphate buffered saline containing 0.1% Tween 20 (PBS-T), the membranes were incubated with mouse anti-His monoclonal antibody (Qiagen).

    Plasmid Preparation:

    Article Title: Natural microbial communities supporting the transfer of the IncP-1β plasmid pB10 exhibit a higher initial content of plasmids from the same incompatibility group
    Article Snippet: .. Basically, sets of qPCR primers were designed to prime on both sides of a unique junction between building blocks of the element targeted for quantification: a unique assembly between two truncated transposons for plasmid pB10 (target coordinates 45968–46102; GenBank NC_004840), and both sides of a unique 97 kb-deletion in the chromosome of E. coli DH5α (target coordinates on K12 274654–372933; GenBank U00096; ; ). qPCRs were performed in triplicate using a “Step One Plus Real-Time PCR System” (Applied Biosystems, driver: StepOne Software v2.2) with thermocycling conditions set as follows: 2 min at 50°C, then 10 min at 95°C followed by 45 cycles of 15 s at 95°C and 1 min at 60°C. .. Quantifications were carried out from 25 ng of community DNA using a “TaqMan® Universal PCR Master Mix, NoAmpErase® UNG” (Applied Biosystems) as recommended by the manufacturer, with 800 nM of each primer, and 300 nM of TaqMan probe, in a 25 μL reaction volume.

    Article Title: Requirement of MrpH for Mannose-Resistant Proteus-Like Fimbria-Mediated Hemagglutination by Proteus mirabilis
    Article Snippet: .. The isopropyl-β- d -thiogalactopyranoside (IPTG)-induced cell lysate (about 10 mg protein) of E. coli DH5α transformed with plasmid pMAL-C2 was coupled to an AminoLink Plus column (Pierce Inc., Rockford, Ill.) as specified by the manufacturer. ..

    Software:

    Article Title: Natural microbial communities supporting the transfer of the IncP-1β plasmid pB10 exhibit a higher initial content of plasmids from the same incompatibility group
    Article Snippet: .. Basically, sets of qPCR primers were designed to prime on both sides of a unique junction between building blocks of the element targeted for quantification: a unique assembly between two truncated transposons for plasmid pB10 (target coordinates 45968–46102; GenBank NC_004840), and both sides of a unique 97 kb-deletion in the chromosome of E. coli DH5α (target coordinates on K12 274654–372933; GenBank U00096; ; ). qPCRs were performed in triplicate using a “Step One Plus Real-Time PCR System” (Applied Biosystems, driver: StepOne Software v2.2) with thermocycling conditions set as follows: 2 min at 50°C, then 10 min at 95°C followed by 45 cycles of 15 s at 95°C and 1 min at 60°C. .. Quantifications were carried out from 25 ng of community DNA using a “TaqMan® Universal PCR Master Mix, NoAmpErase® UNG” (Applied Biosystems) as recommended by the manufacturer, with 800 nM of each primer, and 300 nM of TaqMan probe, in a 25 μL reaction volume.

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    Thermo Fisher e coli dh5α
    PCR amplification of E.coli <t>DH5α</t> clones Lane 2 to 7 –positive amplicons Lane1- DNA marker
    E Coli Dh5α, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 170 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/e coli dh5α/product/Thermo Fisher
    Average 94 stars, based on 170 article reviews
    Price from $9.99 to $1999.99
    e coli dh5α - by Bioz Stars, 2020-07
    94/100 stars
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    93
    Thermo Fisher luciferase expressing e coli dh5α paklux2
    ICE Mh1 PM22 fitness costs, addiction, and consequences for antimicrobial resistance in transconjugants. (A) Growth curves of P. multocida CCUG 17976 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (B) Growth curves of M. haemolytica ATCC 33396 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (C) Luciferase-based competition index from co-cultures of ICE Mh1 PM22 transconjugants and E. coli <t>DH5α</t> harboring the luciferase expression plasmid <t>pAKlux2.</t> Mean of 4 biological replicates with SEM; t-test, ∗ P ≤ 0.05. (D) Long-term repeated passage of ICE Mh1 PM22 transconjugants. Transconjugants were grown in MH (no drug) or MH + 0.5 MIC (subinhibitory; MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396 WT) for 150 days and plated on media with or without selective concentrations of indicated antimicrobials (i.e., 0.5 MIC for non-susceptible isogenic transconjugants). Mean of 3 biological replicates with SEM. (E) Upper panel: growth curves of co-cultures of E. coli DH5α pAKlux2 and E. coli DH5α, P. multocida CCUG 17976 ICE Mh1 PM22 , or M. haemolytica ATCC 33396 ICE Mh1 PM22 in subinhibitory (0.5 MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396) concentrations of oxytetracycline (left) or spectinomycin (right). Mean of 3 biological replicates with SEM. Lower panel: detection of E. coli luciferase production in co-cultures (as above) with either oxytetracycline (left) or spectinomycin (right). (F) Schematic of checkerboard synergy assay for drug interaction screening.
    Luciferase Expressing E Coli Dh5α Paklux2, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/luciferase expressing e coli dh5α paklux2/product/Thermo Fisher
    Average 93 stars, based on 1 article reviews
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    luciferase expressing e coli dh5α paklux2 - by Bioz Stars, 2020-07
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    84
    Thermo Fisher multishot stripwell dh5α t1r competent cells
    Expression of different glycosyl hydrolase (GH) genes from a common expression vector backbone and test of their effects on growth of E. coli <t>DH5α</t> in MOPS minimal medium ( 17 ) and Z. mobilis ZM4 in Zymomonas minimal medium ( 18 ) with glucose or cellobiose as a carbon source. (A) Expression of GH genes in a pVector backbone and a summary of the constructs and their effects on growth in minimal medium supplemented with cellobiose. (B-D) Growth of E. coli DH5α containing plasmids pVector or pCel3A or pGH3 in MOPS minimal medium supplied with 0.4% glucose or cellobiose. (E-G) Growth of Z. mobilis ZM4 containing plasmids pVector or pCel3A or pGH3 in a Zymomonas minimal medium containing 2% glucose or 2% cellobiose. The growth curves are averages of three replicates. *Gene from Cellvibrio japonicus , # Gene from Caulobacter crescentus .
    Multishot Stripwell Dh5α T1r Competent Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    PCR amplification of E.coli DH5α clones Lane 2 to 7 –positive amplicons Lane1- DNA marker

    Journal: Veterinary World

    Article Title: Cloning and sequence analysis of a partial CDS of leptospiral ligA gene in pET-32a – Escherichia coli DH5α system

    doi: 10.14202/vetworld.2018.557-561

    Figure Lengend Snippet: PCR amplification of E.coli DH5α clones Lane 2 to 7 –positive amplicons Lane1- DNA marker

    Article Snippet: The pET-32a ligA plasmid construct was extracted from the transformed E. coli DH5α using Thermo Scientific Gene JET plasmid Miniprep Kit and subjected to sequencing.

    Techniques: Polymerase Chain Reaction, Amplification, Clone Assay, Marker

    PCR amplification of E.coli DH5α clones Lane 2 to 7 –positive amplicons Lane1- DNA marker

    Journal: Veterinary World

    Article Title: Cloning and sequence analysis of a partial CDS of leptospiral ligA gene in pET-32a – Escherichia coli DH5α system

    doi: 10.14202/vetworld.2018.557-561

    Figure Lengend Snippet: PCR amplification of E.coli DH5α clones Lane 2 to 7 –positive amplicons Lane1- DNA marker

    Article Snippet: The pET-32a ligA plasmid construct was extracted from the transformed E. coli DH5α using Thermo Scientific Gene JET plasmid Miniprep Kit and subjected to sequencing.

    Techniques: Polymerase Chain Reaction, Amplification, Clone Assay, Marker

    ICE Mh1 PM22 fitness costs, addiction, and consequences for antimicrobial resistance in transconjugants. (A) Growth curves of P. multocida CCUG 17976 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (B) Growth curves of M. haemolytica ATCC 33396 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (C) Luciferase-based competition index from co-cultures of ICE Mh1 PM22 transconjugants and E. coli DH5α harboring the luciferase expression plasmid pAKlux2. Mean of 4 biological replicates with SEM; t-test, ∗ P ≤ 0.05. (D) Long-term repeated passage of ICE Mh1 PM22 transconjugants. Transconjugants were grown in MH (no drug) or MH + 0.5 MIC (subinhibitory; MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396 WT) for 150 days and plated on media with or without selective concentrations of indicated antimicrobials (i.e., 0.5 MIC for non-susceptible isogenic transconjugants). Mean of 3 biological replicates with SEM. (E) Upper panel: growth curves of co-cultures of E. coli DH5α pAKlux2 and E. coli DH5α, P. multocida CCUG 17976 ICE Mh1 PM22 , or M. haemolytica ATCC 33396 ICE Mh1 PM22 in subinhibitory (0.5 MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396) concentrations of oxytetracycline (left) or spectinomycin (right). Mean of 3 biological replicates with SEM. Lower panel: detection of E. coli luciferase production in co-cultures (as above) with either oxytetracycline (left) or spectinomycin (right). (F) Schematic of checkerboard synergy assay for drug interaction screening.

    Journal: Frontiers in Microbiology

    Article Title: Emerging Variants of the Integrative and Conjugant Element ICEMh1 in Livestock Pathogens: Structural Insights, Potential Host Range, and Implications for Bacterial Fitness and Antimicrobial Therapy

    doi: 10.3389/fmicb.2019.02608

    Figure Lengend Snippet: ICE Mh1 PM22 fitness costs, addiction, and consequences for antimicrobial resistance in transconjugants. (A) Growth curves of P. multocida CCUG 17976 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (B) Growth curves of M. haemolytica ATCC 33396 (spontaneous rifampin-resistant mutant) and isogenic ICE Mh1 PM22 transconjugant. Mean of 4 biological replicates with SEM. (C) Luciferase-based competition index from co-cultures of ICE Mh1 PM22 transconjugants and E. coli DH5α harboring the luciferase expression plasmid pAKlux2. Mean of 4 biological replicates with SEM; t-test, ∗ P ≤ 0.05. (D) Long-term repeated passage of ICE Mh1 PM22 transconjugants. Transconjugants were grown in MH (no drug) or MH + 0.5 MIC (subinhibitory; MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396 WT) for 150 days and plated on media with or without selective concentrations of indicated antimicrobials (i.e., 0.5 MIC for non-susceptible isogenic transconjugants). Mean of 3 biological replicates with SEM. (E) Upper panel: growth curves of co-cultures of E. coli DH5α pAKlux2 and E. coli DH5α, P. multocida CCUG 17976 ICE Mh1 PM22 , or M. haemolytica ATCC 33396 ICE Mh1 PM22 in subinhibitory (0.5 MIC for susceptible P. multocida CCUG 17976 or M. haemolytica ATCC 33396) concentrations of oxytetracycline (left) or spectinomycin (right). Mean of 3 biological replicates with SEM. Lower panel: detection of E. coli luciferase production in co-cultures (as above) with either oxytetracycline (left) or spectinomycin (right). (F) Schematic of checkerboard synergy assay for drug interaction screening.

    Article Snippet: For 2-strain co-culture experiments, luciferase-expressing E. coli DH5α pAKlux2 and test strains (including DH5α without pAKlux2 as a control) were each inoculated at an OD600 of 0.0025 (i.e., total OD600 of ∼0.005) into black clear bottom 96-well plates (Nunc, Thermo-Fisher Scientific, Ottawa, ON, Canada).

    Techniques: Mutagenesis, Luciferase, Expressing, Plasmid Preparation

    Expression of different glycosyl hydrolase (GH) genes from a common expression vector backbone and test of their effects on growth of E. coli DH5α in MOPS minimal medium ( 17 ) and Z. mobilis ZM4 in Zymomonas minimal medium ( 18 ) with glucose or cellobiose as a carbon source. (A) Expression of GH genes in a pVector backbone and a summary of the constructs and their effects on growth in minimal medium supplemented with cellobiose. (B-D) Growth of E. coli DH5α containing plasmids pVector or pCel3A or pGH3 in MOPS minimal medium supplied with 0.4% glucose or cellobiose. (E-G) Growth of Z. mobilis ZM4 containing plasmids pVector or pCel3A or pGH3 in a Zymomonas minimal medium containing 2% glucose or 2% cellobiose. The growth curves are averages of three replicates. *Gene from Cellvibrio japonicus , # Gene from Caulobacter crescentus .

    Journal: bioRxiv

    Article Title: Heterologous glycosyl hydrolase expression and cellular reprogramming resembling sucrose-induction enable Zymomonas mobilis growth on cellobiose

    doi: 10.1101/854646

    Figure Lengend Snippet: Expression of different glycosyl hydrolase (GH) genes from a common expression vector backbone and test of their effects on growth of E. coli DH5α in MOPS minimal medium ( 17 ) and Z. mobilis ZM4 in Zymomonas minimal medium ( 18 ) with glucose or cellobiose as a carbon source. (A) Expression of GH genes in a pVector backbone and a summary of the constructs and their effects on growth in minimal medium supplemented with cellobiose. (B-D) Growth of E. coli DH5α containing plasmids pVector or pCel3A or pGH3 in MOPS minimal medium supplied with 0.4% glucose or cellobiose. (E-G) Growth of Z. mobilis ZM4 containing plasmids pVector or pCel3A or pGH3 in a Zymomonas minimal medium containing 2% glucose or 2% cellobiose. The growth curves are averages of three replicates. *Gene from Cellvibrio japonicus , # Gene from Caulobacter crescentus .

    Article Snippet: The E. coli DH10B strain was used for cloning and E. coli DH5α was used for expressing the recombinant plasmids.

    Techniques: Expressing, Plasmid Preparation, Construct