Structured Review

Difco middlebrook 7h9 broth
Sources of RNA samples. ( A ) MTB H37Rv was grown in <t>Middlebrook</t> <t>7H9</t> medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.
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1) Product Images from "Differential expression of iron-, carbon-, and oxygen-responsive mycobacterial genes in the lungs of chronically infected mice and tuberculosis patients"

Article Title: Differential expression of iron-, carbon-, and oxygen-responsive mycobacterial genes in the lungs of chronically infected mice and tuberculosis patients

Journal: Proceedings of the National Academy of Sciences of the United States of America

doi: 10.1073/pnas.2436197100

Sources of RNA samples. ( A ) MTB H37Rv was grown in Middlebrook 7H9 medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.
Figure Legend Snippet: Sources of RNA samples. ( A ) MTB H37Rv was grown in Middlebrook 7H9 medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.

Techniques Used: RNA Extraction, Quantitative RT-PCR, Mouse Assay, Infection, Staining

2) Product Images from "Organic Hydroperoxide Resistance Protein and Ergothioneine Compensate for Loss of Mycothiol in Mycobacterium smegmatis Mutants ▿ Mutants ▿ †"

Article Title: Organic Hydroperoxide Resistance Protein and Ergothioneine Compensate for Loss of Mycothiol in Mycobacterium smegmatis Mutants ▿ Mutants ▿ †

Journal: Journal of Bacteriology

doi: 10.1128/JB.01402-10

(A) Resistance of M. smegmatis to CuOOH is dependent in part upon Ohr. Cells were cultured to late stationary phase (17 days) in Middlebrook 7H9 medium. Cells were then suspended in the same medium at an A 600 of 0.1 and incubated with CuOOH for 2 h at
Figure Legend Snippet: (A) Resistance of M. smegmatis to CuOOH is dependent in part upon Ohr. Cells were cultured to late stationary phase (17 days) in Middlebrook 7H9 medium. Cells were then suspended in the same medium at an A 600 of 0.1 and incubated with CuOOH for 2 h at

Techniques Used: Cell Culture, Incubation

3) Product Images from "Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk"

Article Title: Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk

Journal: Applied and Environmental Microbiology

doi: 10.1128/AEM.71.6.2853-2861.2005

Clump size distributions in Middlebrook 7H9 broth suspensions of three M. avium subsp. paratuberculosis strains determined using Mastersizer.
Figure Legend Snippet: Clump size distributions in Middlebrook 7H9 broth suspensions of three M. avium subsp. paratuberculosis strains determined using Mastersizer.

Techniques Used:

Clump size distribution in (A) an untreated Middlebrook 7H9 broth suspension of M. avium subsp. paratuberculosis NCTC 8578, (B) the same broth culture homogenized at 2,500 lb/in 2 and analyzed immediately, and (C) the same broth culture analyzed 10 min
Figure Legend Snippet: Clump size distribution in (A) an untreated Middlebrook 7H9 broth suspension of M. avium subsp. paratuberculosis NCTC 8578, (B) the same broth culture homogenized at 2,500 lb/in 2 and analyzed immediately, and (C) the same broth culture analyzed 10 min

Techniques Used:

4) Product Images from "Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿"

Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿

Journal: Applied and Environmental Microbiology

doi: 10.1128/AEM.00981-08

Effects of different concentrations (μg/ml) of trans -cinnamaldehyde and noninhibitory concentrations of ethanol (0.4%) (negative control) on the growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth.
Figure Legend Snippet: Effects of different concentrations (μg/ml) of trans -cinnamaldehyde and noninhibitory concentrations of ethanol (0.4%) (negative control) on the growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth.

Techniques Used: Negative Control

Growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth supplemented with the six active compounds at their MICs ( trans -cinnamaldehyde [25.9 μg/ml], cinnamon oil [26.2 μg/ml], carvacrol [72.2 μg/ml], oregano oil [68.2 μg/ml], 2-hydroxy-5-methoxybenzaldehyde [90.4 μg/ml], and 2,5-dihydroxybenzaldehyde [74 μg/ml]), ethanol (0.4%) (negative control), vanillic acid (74 μg/ml), and citral (65.7 μg/ml). 2-Hydroxy-5-methoxybenzaldehyde and 2,5-dihydroxybenzaldehyde had slightly higher initial OD values as they produced colored solutions.
Figure Legend Snippet: Growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth supplemented with the six active compounds at their MICs ( trans -cinnamaldehyde [25.9 μg/ml], cinnamon oil [26.2 μg/ml], carvacrol [72.2 μg/ml], oregano oil [68.2 μg/ml], 2-hydroxy-5-methoxybenzaldehyde [90.4 μg/ml], and 2,5-dihydroxybenzaldehyde [74 μg/ml]), ethanol (0.4%) (negative control), vanillic acid (74 μg/ml), and citral (65.7 μg/ml). 2-Hydroxy-5-methoxybenzaldehyde and 2,5-dihydroxybenzaldehyde had slightly higher initial OD values as they produced colored solutions.

Techniques Used: Negative Control, Produced

5) Product Images from "LprG-Mediated Surface Expression of Lipoarabinomannan Is Essential for Virulence of Mycobacterium tuberculosis"

Article Title: LprG-Mediated Surface Expression of Lipoarabinomannan Is Essential for Virulence of Mycobacterium tuberculosis

Journal: PLoS Pathogens

doi: 10.1371/journal.ppat.1004376

The lprG mutant is attenuated in mouse macrophages. Resting and IFN-γ-activated BMMΦ from C57BL/6 (A, B, E, and F), p-47phox−/− (C and D), and iNOS−/− (G, H) mice were infected with wild-type (H37Rv), lprG mutant (Δ lprG ), and Δ lprG complemented with lprG -Rv1410c (:: lprG ) at MOI of 1∶10 (A–D) and 1∶40 (E–H). Intracellular bacteria were enumerated on Middlebrook 7H9 agar on the indicated days. Data points represent average CFU ±SD of triplicate wells. Data is representative of two to three independent experiments.
Figure Legend Snippet: The lprG mutant is attenuated in mouse macrophages. Resting and IFN-γ-activated BMMΦ from C57BL/6 (A, B, E, and F), p-47phox−/− (C and D), and iNOS−/− (G, H) mice were infected with wild-type (H37Rv), lprG mutant (Δ lprG ), and Δ lprG complemented with lprG -Rv1410c (:: lprG ) at MOI of 1∶10 (A–D) and 1∶40 (E–H). Intracellular bacteria were enumerated on Middlebrook 7H9 agar on the indicated days. Data points represent average CFU ±SD of triplicate wells. Data is representative of two to three independent experiments.

Techniques Used: Mutagenesis, Mouse Assay, Infection

6) Product Images from "Rapid Colorimetric Method for Testing Susceptibility of Mycobacterium tuberculosis to Isoniazid and Rifampin in Liquid Cultures"

Article Title: Rapid Colorimetric Method for Testing Susceptibility of Mycobacterium tuberculosis to Isoniazid and Rifampin in Liquid Cultures

Journal: Journal of Clinical Microbiology

doi: 10.1128/JCM.41.11.5173-5177.2003

Nitrite production ( y axis) measured in 24 M. tuberculosis strains during 2 weeks' growth in Middlebrook 7H9 broth with supplements.
Figure Legend Snippet: Nitrite production ( y axis) measured in 24 M. tuberculosis strains during 2 weeks' growth in Middlebrook 7H9 broth with supplements.

Techniques Used:

7) Product Images from "Mycobacterium tuberculosis CDC1551 Is Resistant to Reactive Nitrogen and Oxygen Intermediates In Vitro "

Article Title: Mycobacterium tuberculosis CDC1551 Is Resistant to Reactive Nitrogen and Oxygen Intermediates In Vitro

Journal: Infection and Immunity

doi: 10.1128/IAI.70.7.3965-3968.2002

Effect of H 2 O 2 on mycobacterial viability in Middlebrook 7H9 medium at 2 mM (black bars) and 5 mM (gray bars) concentrations. Data shown are representative of triplicate experiments. Results are expressed as percents survival relative to baseline ± standard errors of the means and are mean values of triplicate cultures for each strain.
Figure Legend Snippet: Effect of H 2 O 2 on mycobacterial viability in Middlebrook 7H9 medium at 2 mM (black bars) and 5 mM (gray bars) concentrations. Data shown are representative of triplicate experiments. Results are expressed as percents survival relative to baseline ± standard errors of the means and are mean values of triplicate cultures for each strain.

Techniques Used:

8) Product Images from "Development of a repressible mycobacterial promoter system based on two transcriptional repressors"

Article Title: Development of a repressible mycobacterial promoter system based on two transcriptional repressors

Journal: Nucleic Acids Research

doi: 10.1093/nar/gkq235

Growth curve of the fadD32 conditional mutant TB47 in the presence of ATc and pristinamycin I. Bacteria were grown in Middlebrook 7H9 without ATc (diamonds) or with 200 ng/ml ATc (all the others). Forty-eight hours after the beginning of the experiment cultures were supplemented with 20 (filled triangles), 200 (circles) or 2000 ng/ml (open triangles) of pristinamycin I. As a control, one culture grown in 200 ng/ml ATc did not receive pristinamycin I (squares).
Figure Legend Snippet: Growth curve of the fadD32 conditional mutant TB47 in the presence of ATc and pristinamycin I. Bacteria were grown in Middlebrook 7H9 without ATc (diamonds) or with 200 ng/ml ATc (all the others). Forty-eight hours after the beginning of the experiment cultures were supplemented with 20 (filled triangles), 200 (circles) or 2000 ng/ml (open triangles) of pristinamycin I. As a control, one culture grown in 200 ng/ml ATc did not receive pristinamycin I (squares).

Techniques Used: Mutagenesis

Characterization of the M. smegmatis ftsZ mutant MS98. ( A ) MS98 was plated on Middlebrook 7H10 medium with or without 50 ng/ml ATc. ( B ) MS98 and its parental wild-type strain were grown in Middlebrook 7H9 with or without ATc.
Figure Legend Snippet: Characterization of the M. smegmatis ftsZ mutant MS98. ( A ) MS98 was plated on Middlebrook 7H10 medium with or without 50 ng/ml ATc. ( B ) MS98 and its parental wild-type strain were grown in Middlebrook 7H9 with or without ATc.

Techniques Used: Mutagenesis

β-Galactosidase assay in liquid media. ( A ) M. smegmatis MS82 was pre-grown in Middlebrook 7H9 containing 50 ng/ml ATc, and diluted into fresh media containing different ATc concentrations. Every 24 h and for 3 days, one aliquot of each culture was used to measure β-galactosidase activity and one aliquot was diluted into fresh medium containing the same amount of ATc as before. Black bars: 24 h; gray bar: 48 h; white bars: 72 h. ( B ) TB38.2 was pre-grown in Middlebrook 7H9 containing 200 ng/ml ATc, and diluted into fresh media containing different ATc concentrations. Every 48 h and for 6 days, one aliquot of each culture was used to measure β-galactosidase activity and one aliquot was diluted into fresh medium containing the same amount of ATc as before. Black bars: 48 h; grey bar: 96 h; white bars: 144 h. Values are indicated as percentages of maximal activity.
Figure Legend Snippet: β-Galactosidase assay in liquid media. ( A ) M. smegmatis MS82 was pre-grown in Middlebrook 7H9 containing 50 ng/ml ATc, and diluted into fresh media containing different ATc concentrations. Every 24 h and for 3 days, one aliquot of each culture was used to measure β-galactosidase activity and one aliquot was diluted into fresh medium containing the same amount of ATc as before. Black bars: 24 h; gray bar: 48 h; white bars: 72 h. ( B ) TB38.2 was pre-grown in Middlebrook 7H9 containing 200 ng/ml ATc, and diluted into fresh media containing different ATc concentrations. Every 48 h and for 6 days, one aliquot of each culture was used to measure β-galactosidase activity and one aliquot was diluted into fresh medium containing the same amount of ATc as before. Black bars: 48 h; grey bar: 96 h; white bars: 144 h. Values are indicated as percentages of maximal activity.

Techniques Used: Activity Assay

Characterization of the TetR/Pip OFF system. ( A ) β-galactosidase assay in M. smegmatis in liquid media: MS82 and MS83 were grown in Middlebrook 7H9 medium with or without 50 ng/ml ATc. White bars: bacteria grown without ATc; grey bars: bacteria exposed to ATc for 6 h; black bars: bacteria exposed to ATc for 18 h. ( B ) β-galactosidase assay in M. tuberculosis in liquid media: TB38.1 and TB38.2 were grown in Middlebrook 7H9 medium with or without 200 ng/ml ATc. White bars: bacteria grown without ATc; black bars: bacteria exposed to ATc for 48 h. Results are expressed in Miller units.
Figure Legend Snippet: Characterization of the TetR/Pip OFF system. ( A ) β-galactosidase assay in M. smegmatis in liquid media: MS82 and MS83 were grown in Middlebrook 7H9 medium with or without 50 ng/ml ATc. White bars: bacteria grown without ATc; grey bars: bacteria exposed to ATc for 6 h; black bars: bacteria exposed to ATc for 18 h. ( B ) β-galactosidase assay in M. tuberculosis in liquid media: TB38.1 and TB38.2 were grown in Middlebrook 7H9 medium with or without 200 ng/ml ATc. White bars: bacteria grown without ATc; black bars: bacteria exposed to ATc for 48 h. Results are expressed in Miller units.

Techniques Used:

Characterization of the M. tuberculosis fadD32 conditional mutant TB47. ( A ) Growth curve in the presence of different concentrations of ATc. TB47 was grown in Middlebrook 7H9 medium containing 200 ng/ml ATc (empty triangles), 100 ng/ml ATc (crosses), 50 ng/ml ATc (filled triangles) or No ATc (empty squares). ( B ) Viable counts variation during growth inhibition due to fadD32 depletion. TB47 was grown in Middlebrook 7H9 medium containing 200 ng/ml ATc. An aliquot of the bacterial culture was collected at different time points starting from 48 h after the beginning of the experiment, diluted and plated for cfu determination. Squares: cfu/ml; diamonds:optical density at 540 nm.
Figure Legend Snippet: Characterization of the M. tuberculosis fadD32 conditional mutant TB47. ( A ) Growth curve in the presence of different concentrations of ATc. TB47 was grown in Middlebrook 7H9 medium containing 200 ng/ml ATc (empty triangles), 100 ng/ml ATc (crosses), 50 ng/ml ATc (filled triangles) or No ATc (empty squares). ( B ) Viable counts variation during growth inhibition due to fadD32 depletion. TB47 was grown in Middlebrook 7H9 medium containing 200 ng/ml ATc. An aliquot of the bacterial culture was collected at different time points starting from 48 h after the beginning of the experiment, diluted and plated for cfu determination. Squares: cfu/ml; diamonds:optical density at 540 nm.

Techniques Used: Mutagenesis, Inhibition

9) Product Images from "Deficiency of the Novel Exopolyphosphatase Rv1026/PPX2 Leads to Metabolic Downshift and Altered Cell Wall Permeability in Mycobacterium tuberculosis"

Article Title: Deficiency of the Novel Exopolyphosphatase Rv1026/PPX2 Leads to Metabolic Downshift and Altered Cell Wall Permeability in Mycobacterium tuberculosis

Journal: mBio

doi: 10.1128/mBio.02428-14

Intrabacillary accumulation of inorganic polyphosphate temporally coincides with M. tuberculosis growth restriction. (A to D) Intrabacillary poly(P) content was measured in wild-type M. tuberculosis during axenic growth in supplemented Middlebrook 7H9 broth (A), nutrient starvation (B), progressive hypoxia (C), and phosphate depletion (D) using a DAPI-based method and normalized to total protein content of extract lysate. Each data point represents the mean of three biological replicates. In the progressive hypoxia model in panel C, the x axis shows the days after change in color of the indicator dye, methylene blue, indicating bacterial entry into nonreplicating persistence stage 2 (mean ± SD). In panels A to D, the poly(P)/total protein ratio is shown on the left-hand x axes, and the optical density at 600 nm (O.D. 600) is shown on the right-hand x axes.
Figure Legend Snippet: Intrabacillary accumulation of inorganic polyphosphate temporally coincides with M. tuberculosis growth restriction. (A to D) Intrabacillary poly(P) content was measured in wild-type M. tuberculosis during axenic growth in supplemented Middlebrook 7H9 broth (A), nutrient starvation (B), progressive hypoxia (C), and phosphate depletion (D) using a DAPI-based method and normalized to total protein content of extract lysate. Each data point represents the mean of three biological replicates. In the progressive hypoxia model in panel C, the x axis shows the days after change in color of the indicator dye, methylene blue, indicating bacterial entry into nonreplicating persistence stage 2 (mean ± SD). In panels A to D, the poly(P)/total protein ratio is shown on the left-hand x axes, and the optical density at 600 nm (O.D. 600) is shown on the right-hand x axes.

Techniques Used:

10) Product Images from "Novel Method for Rapid Measurement of Growth of Mycobacteria in Detergent-Free Media"

Article Title: Novel Method for Rapid Measurement of Growth of Mycobacteria in Detergent-Free Media

Journal: Journal of Clinical Microbiology

doi:

Growth curves of M. tuberculosis H37Rv (I) and M. bovis BCG Tokyo (II) cultures in Middlebrook 7H9 broth. Duplicate cultures were grown with (•, ○) and without (▾, ▿) 0.05% Tween 80 and supplemented with ADC ( M. tuberculosis H37Rv) or OADC ( M. bovis BCG Tokyo). Curves were plotted from total protein yield (a), OD (b), and total ATP (c) data. The dashed horizontal lines on the protein growth curves indicate the lower limit of sensitivity of the protein assay (6 μg of protein/ml). Each data point represents the average of two measurements. Error bars show the standard deviations of the means.
Figure Legend Snippet: Growth curves of M. tuberculosis H37Rv (I) and M. bovis BCG Tokyo (II) cultures in Middlebrook 7H9 broth. Duplicate cultures were grown with (•, ○) and without (▾, ▿) 0.05% Tween 80 and supplemented with ADC ( M. tuberculosis H37Rv) or OADC ( M. bovis BCG Tokyo). Curves were plotted from total protein yield (a), OD (b), and total ATP (c) data. The dashed horizontal lines on the protein growth curves indicate the lower limit of sensitivity of the protein assay (6 μg of protein/ml). Each data point represents the average of two measurements. Error bars show the standard deviations of the means.

Techniques Used:

11) Product Images from "LipG a bifunctional phospholipase/thioesterase involved in mycobacterial envelope remodeling"

Article Title: LipG a bifunctional phospholipase/thioesterase involved in mycobacterial envelope remodeling

Journal: Bioscience Reports

doi: 10.1042/BSR20181953

Disruption of lipG MS triggers cell-wall modifications and decreases antibiotics susceptibility ( A ) Colony morphology of WT, lipG MS disrupted and complemented strains. Recombinant strains were grown in 7H9-S before being spotted on to Middlebrook 7H11 agar plate devoid of Tween-80. Images are representative of two independent experiments. ( B ) GPL analysis of WT, lipG MS disrupted and complemented strains. Cells were grown in 7H9-S until OD 600 nm ∼1–1.5. Lipid extraction was followed by TLC (left panel) and densitometry analysis (right panel). Results are expressed as mean ± S.D. of total GPL level from three independent experiments, where WT level was arbitrarily adjusted to 1 relative unit. Statistical analysis was performed by using the Student’s t test. Relative GPL level from lipG MS disrupted mutant were compared with the WT where * corresponded to a P -value
Figure Legend Snippet: Disruption of lipG MS triggers cell-wall modifications and decreases antibiotics susceptibility ( A ) Colony morphology of WT, lipG MS disrupted and complemented strains. Recombinant strains were grown in 7H9-S before being spotted on to Middlebrook 7H11 agar plate devoid of Tween-80. Images are representative of two independent experiments. ( B ) GPL analysis of WT, lipG MS disrupted and complemented strains. Cells were grown in 7H9-S until OD 600 nm ∼1–1.5. Lipid extraction was followed by TLC (left panel) and densitometry analysis (right panel). Results are expressed as mean ± S.D. of total GPL level from three independent experiments, where WT level was arbitrarily adjusted to 1 relative unit. Statistical analysis was performed by using the Student’s t test. Relative GPL level from lipG MS disrupted mutant were compared with the WT where * corresponded to a P -value

Techniques Used: Mass Spectrometry, Recombinant, Thin Layer Chromatography, Mutagenesis

12) Product Images from "Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis"

Article Title: Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis

Journal: Infection and Immunity

doi:

Inactivation of leuD confers leucine auxotrophy. Bacterial growth in Middlebrook 7H9 broth with and without supplementation with 50 μg of leucine per ml. The various strains were cultured in 7H9 medium supplemented with leucine and then pelleted, washed, and resuspended in media with and without leucine supplementation. The OD 600 s of the broth cultures were determined daily.
Figure Legend Snippet: Inactivation of leuD confers leucine auxotrophy. Bacterial growth in Middlebrook 7H9 broth with and without supplementation with 50 μg of leucine per ml. The various strains were cultured in 7H9 medium supplemented with leucine and then pelleted, washed, and resuspended in media with and without leucine supplementation. The OD 600 s of the broth cultures were determined daily.

Techniques Used: Cell Culture

13) Product Images from "Mimicry of the Pathogenic Mycobacterium Vacuole In Vitro Elicits the Bacterial Intracellular Phenotype, Including Early-Onset Macrophage Death ▿"

Article Title: Mimicry of the Pathogenic Mycobacterium Vacuole In Vitro Elicits the Bacterial Intracellular Phenotype, Including Early-Onset Macrophage Death ▿

Journal: Infection and Immunity

doi: 10.1128/IAI.01120-10

Real-time PCR of MAC strain 104 incubated in single metals. MAC was exposed to Middlebrook 7H9 broth containing the indicated volume of a 1 M concentration of the indicated metal per 500 ml of Middlebrook 7H9 broth for 24 h, and then RNA was extracted
Figure Legend Snippet: Real-time PCR of MAC strain 104 incubated in single metals. MAC was exposed to Middlebrook 7H9 broth containing the indicated volume of a 1 M concentration of the indicated metal per 500 ml of Middlebrook 7H9 broth for 24 h, and then RNA was extracted

Techniques Used: Real-time Polymerase Chain Reaction, Incubation, Concentration Assay

Uptake of Mycobacterium avium complex (MAC) incubated in the elemental mixture by macrophages. (A) MAC was incubated in Middlebrook 7H9 broth, the 1-h elemental mixture, and the 24-h elemental mixture for 1 h and 24 h, respectively, and then used to infect
Figure Legend Snippet: Uptake of Mycobacterium avium complex (MAC) incubated in the elemental mixture by macrophages. (A) MAC was incubated in Middlebrook 7H9 broth, the 1-h elemental mixture, and the 24-h elemental mixture for 1 h and 24 h, respectively, and then used to infect

Techniques Used: Incubation

Real-time PCR of MAC strain 104 in the subtracted 24-h elemental mixtures. Total bacterial RNA was extracted from bacteria exposed to the indicated mixture for 24 h and bacteria exposed to Middlebrook 7H9 broth as a reference. cDNA was made from the RNA
Figure Legend Snippet: Real-time PCR of MAC strain 104 in the subtracted 24-h elemental mixtures. Total bacterial RNA was extracted from bacteria exposed to the indicated mixture for 24 h and bacteria exposed to Middlebrook 7H9 broth as a reference. cDNA was made from the RNA

Techniques Used: Real-time Polymerase Chain Reaction

Real-time PCR of MAC strain 104 exposed to the 24-h elemental mixture. Total bacterial RNA was extracted from MAC strain 104 exposed to the 24-h elemental mixture for 24 h and bacteria exposed to Middlebrook 7H9 broth as a control. cDNA was made from
Figure Legend Snippet: Real-time PCR of MAC strain 104 exposed to the 24-h elemental mixture. Total bacterial RNA was extracted from MAC strain 104 exposed to the 24-h elemental mixture for 24 h and bacteria exposed to Middlebrook 7H9 broth as a control. cDNA was made from

Techniques Used: Real-time Polymerase Chain Reaction

14) Product Images from "Reversible Lipid Accumulation and Associated Division Arrest of Mycobacterium avium in Lipoprotein-Induced Foamy Macrophages May Resemble Key Events during Latency and Reactivation of Tuberculosis"

Article Title: Reversible Lipid Accumulation and Associated Division Arrest of Mycobacterium avium in Lipoprotein-Induced Foamy Macrophages May Resemble Key Events during Latency and Reactivation of Tuberculosis

Journal: Infection and Immunity

doi: 10.1128/IAI.01196-13

Fatty acids for LB and ILI formation are provided through TAG (from VLDL) breakdown in host lysosomes. (A and B) M. avium cultured in vitro in Middlebrook 7H9 medium for 4 days was exposed to VLDL for 24 h. Bacteria were processed for EM and examined
Figure Legend Snippet: Fatty acids for LB and ILI formation are provided through TAG (from VLDL) breakdown in host lysosomes. (A and B) M. avium cultured in vitro in Middlebrook 7H9 medium for 4 days was exposed to VLDL for 24 h. Bacteria were processed for EM and examined

Techniques Used: Cell Culture, In Vitro

15) Product Images from "senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence"

Article Title: senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence

Journal: BMC Microbiology

doi: 10.1186/s12866-014-0265-8

Gene expression and regulation of senX3 , regX3, and the phosphate-specific transport gene pstC2 in each strain by qRT-PCR. An equal amount of total RNA samples were used for cDNA synthesis with random primers. The cycle threshold value (C T ) of each gene of interest was normalized to that of the housekeeping gene sigA to generate ∆C T , which was then converted to fold change (1.83e-∆C T ). (A) Expression of the genes senX3 and regX3 in Middlebrook 7H9 broth. (B) Expression of the genes senX3 and regX3 during P i depletion. (C) Expression of the pstC2 gene in Middlebrook 7H9 broth. (D) Expression of the pstC2 gene during P i depletion. The samples were prepared as duplicates and the experiment was repeated twice with similar results. * p
Figure Legend Snippet: Gene expression and regulation of senX3 , regX3, and the phosphate-specific transport gene pstC2 in each strain by qRT-PCR. An equal amount of total RNA samples were used for cDNA synthesis with random primers. The cycle threshold value (C T ) of each gene of interest was normalized to that of the housekeeping gene sigA to generate ∆C T , which was then converted to fold change (1.83e-∆C T ). (A) Expression of the genes senX3 and regX3 in Middlebrook 7H9 broth. (B) Expression of the genes senX3 and regX3 during P i depletion. (C) Expression of the pstC2 gene in Middlebrook 7H9 broth. (D) Expression of the pstC2 gene during P i depletion. The samples were prepared as duplicates and the experiment was repeated twice with similar results. * p

Techniques Used: Expressing, Quantitative RT-PCR

16) Product Images from "Altered Mycobacterium tuberculosis Cell Wall Metabolism and Physiology Associated With RpoB Mutation H526D"

Article Title: Altered Mycobacterium tuberculosis Cell Wall Metabolism and Physiology Associated With RpoB Mutation H526D

Journal: Frontiers in Microbiology

doi: 10.3389/fmicb.2018.00494

The Mycobacterium tuberculosis (Mtb) RpoB H526D mutant strain exhibits slow growth rate and altered colony morphology. (A) Growth kinetics of Mtb strains in supplemented Middlebrook 7H9 broth. ∗∗ p
Figure Legend Snippet: The Mycobacterium tuberculosis (Mtb) RpoB H526D mutant strain exhibits slow growth rate and altered colony morphology. (A) Growth kinetics of Mtb strains in supplemented Middlebrook 7H9 broth. ∗∗ p

Techniques Used: Mutagenesis

Related Articles

Clone Assay:

Article Title: Organic Hydroperoxide Resistance Protein and Ergothioneine Compensate for Loss of Mycothiol in Mycobacterium smegmatis Mutants ▿ Mutants ▿ †
Article Snippet: M. smegmatis mc2 155 was grown in Middlebrook 7H9 broth (Difco) with 0.05% Tween 80 and supplemented with either OADC (oleic acid, albumin, glucose, and catalase supplement) or 1% glucose. .. Escherichia coli DH5α was used as the host strain for cloning experiments.

Incubation:

Article Title: Altered Mycobacterium tuberculosis Cell Wall Metabolism and Physiology Associated With RpoB Mutation H526D
Article Snippet: Nutrient-rich conditions were established in Middlebrook 7H9 broth (Difco, Sparks, MD, United States) supplemented with 10% oleic acid-albumin-dextrose-catalase (OADC) (Difco), 0.1% glycerol, and 0.05% Tween-80 at 37°C on a shaker. .. For nutrient starvation experiments, bacterial pellets were washed three times with 1xPBS (Biological Quality) containing 0.05% Tween-80 and re-suspended in 10 ml of the same medium (at OD600 ∼ 0.1) in 50-ml conical tubes prior to standing incubation at 37°C for 14 days ( ).

Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿
Article Snippet: .. Two cryobeads were inoculated into 10 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, MI) supplemented with 0.05% (wt/vol) Tween 80 (Sigma), 10% (vol/vol) Middlebrook oleic acid-albumin-dextrose-catalase supplement (Difco), and 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), pH 6.6 ± 0.2, and the broths were incubated at 37°C with gentle shaking (100 rpm) for 3 to 4 weeks until the optical density at 600 nm (OD600 ) was between 0.7 and 0.9. .. The M. avium subsp. paratuberculosis culture was declumped by adding 10 sterile 3-mm glass beads and then vortexing the culture at high speed for 2 min, resting it for 2 min, and repeating the cycle three times.

Concentration Assay:

Article Title: LipG a bifunctional phospholipase/thioesterase involved in mycobacterial envelope remodeling
Article Snippet: M. smegmatis mc2 155 and M. smegmatis mc2 155 groEL1ΔC [ ] strains were cultured in Middlebrook 7H9 broth (BD Difco, Le Pont-de-Claix, France) supplemented with 0.05% (v/v ) Tween-80 and 0.2% (v/v ) glycerol (Sigma–Aldrich, Saint-Quentin Fallavier, France) (7H9-S). .. Hygromycin B (Euromedex) was used at a final concentration of 200 and 50 µg/ml for recombinant E. coli and recombinant mycobacteria, respectively.

Mutagenesis:

Article Title: Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis
Article Snippet: Liquid cultures of M. tuberculosis H37Rv, M. tuberculosis Erdman, and M. bovis BCG strains Pasteur (BCG-P), mc2 3034, and mc2 3035 were grown in Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) supplemented with 0.2% glycerol, 0.05% Tween 80, and 10% Middlebrook OADC (Difco) (minimal medium). .. This mutant strain is independent of a previously isolated leucine auxotroph created by Balasubramanian and colleagues ( ).

Isolation:

Article Title: Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis
Article Snippet: Liquid cultures of M. tuberculosis H37Rv, M. tuberculosis Erdman, and M. bovis BCG strains Pasteur (BCG-P), mc2 3034, and mc2 3035 were grown in Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) supplemented with 0.2% glycerol, 0.05% Tween 80, and 10% Middlebrook OADC (Difco) (minimal medium). .. This mutant strain is independent of a previously isolated leucine auxotroph created by Balasubramanian and colleagues ( ).

Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿
Article Snippet: 806R was isolated from raw cow's milk by Grant et al. ( ). .. Two cryobeads were inoculated into 10 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, MI) supplemented with 0.05% (wt/vol) Tween 80 (Sigma), 10% (vol/vol) Middlebrook oleic acid-albumin-dextrose-catalase supplement (Difco), and 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), pH 6.6 ± 0.2, and the broths were incubated at 37°C with gentle shaking (100 rpm) for 3 to 4 weeks until the optical density at 600 nm (OD600 ) was between 0.7 and 0.9.

Northern Blot:

Article Title: Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk
Article Snippet: For experiment 5, three other M. avium subsp. paratuberculosis strains were employed: 806R, an M. avium subsp. paratuberculosis isolate from raw milk; B4, an M. avium subsp. paratuberculosis isolate from a bovine in Northern Ireland; and 796PSS, an M. avium subsp. paratuberculosis isolate from pasteurized milk. .. Inoculum for the pilot plant runs was prepared at QUB by inoculating 50 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) containing 10% OADC supplement (Difco), 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), and 0.05% Tween 80 (Sigma Chemical Co. Ltd., Poole, England) with a colony of the appropriate strain of M. avium subsp. paratuberculosis from a Herrold's egg yolk medium (HEYM) slope culture and incubating for 8 weeks at 37°C.

Protein Extraction:

Article Title: Novel Method for Rapid Measurement of Growth of Mycobacteria in Detergent-Free Media
Article Snippet: We wished to monitor the growth of mycobacteria in a BSA-free, Tween-free defined medium and describe here the development of a rapid and reliable protein extraction method for measuring mycobacterial growth in these heavily clumped cultures. (Portions of this work have been presented at the TB: Molecular Mechanisms and Immunologic Aspects, Keystone, Colorado, 1998, meeting.) .. Cultures of M. tuberculosis H37Rv, Mycobacterium bovis BCG (Tokyo), and Mycobacterium smegmatis mc2 155 ( ) were grown with agitation at 37°C in 400 ml of Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.), containing sterile OADC (0.5% BSA, 0.2% glucose, 0.006% oleic acid, 140 mM NaCl) or ADC (0.5% BSA, 0.2% glucose, 3 μg of catalase per ml) supplement as indicated in 1-liter screw-cap bottles.

Cell Culture:

Article Title: LipG a bifunctional phospholipase/thioesterase involved in mycobacterial envelope remodeling
Article Snippet: .. M. smegmatis mc2 155 and M. smegmatis mc2 155 groEL1ΔC [ ] strains were cultured in Middlebrook 7H9 broth (BD Difco, Le Pont-de-Claix, France) supplemented with 0.05% (v/v ) Tween-80 and 0.2% (v/v ) glycerol (Sigma–Aldrich, Saint-Quentin Fallavier, France) (7H9-S). .. When needed, ampicillin and kanamycin (Euromedex, Souffelweyersheim, France) were added to the medium at final concentrations of 100 and 50 µg/ml for both E. coli and mycobacterial species, respectively.

Article Title: Attenuation of and Protection Induced by a Leucine Auxotroph of Mycobacterium tuberculosis
Article Snippet: Liquid cultures of M. tuberculosis H37Rv, M. tuberculosis Erdman, and M. bovis BCG strains Pasteur (BCG-P), mc2 3034, and mc2 3035 were grown in Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) supplemented with 0.2% glycerol, 0.05% Tween 80, and 10% Middlebrook OADC (Difco) (minimal medium). .. Strain mc2 3032 (H37Rv Δ leuD ) was cultured on minimal medium supplemented with 50 μg of l -leucine (Sigma Chemical Co., St. Louis, Mo.)/ml (complete medium).

Drug Susceptibility Assay:

Article Title: Altered Mycobacterium tuberculosis Cell Wall Metabolism and Physiology Associated With RpoB Mutation H526D
Article Snippet: A wild-type Mtb CDC1551 and its lab-derived RpoB mutants H526D ( ) and D516V were also used to confirm our findings only in the vancomycin susceptibility assay. .. Nutrient-rich conditions were established in Middlebrook 7H9 broth (Difco, Sparks, MD, United States) supplemented with 10% oleic acid-albumin-dextrose-catalase (OADC) (Difco), 0.1% glycerol, and 0.05% Tween-80 at 37°C on a shaker.

Quantitation Assay:

Article Title: Novel Method for Rapid Measurement of Growth of Mycobacteria in Detergent-Free Media
Article Snippet: Although the BACTEC system is widely used in clinical laboratories for the radiometric quantitation of mycobacterial growth , it is inflexible in that it is limited to a single, prepacked growth medium (Middlebrook 7H12 broth). .. Cultures of M. tuberculosis H37Rv, Mycobacterium bovis BCG (Tokyo), and Mycobacterium smegmatis mc2 155 ( ) were grown with agitation at 37°C in 400 ml of Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.), containing sterile OADC (0.5% BSA, 0.2% glucose, 0.006% oleic acid, 140 mM NaCl) or ADC (0.5% BSA, 0.2% glucose, 3 μg of catalase per ml) supplement as indicated in 1-liter screw-cap bottles.

other:

Article Title: Deficiency of the Novel Exopolyphosphatase Rv1026/PPX2 Leads to Metabolic Downshift and Altered Cell Wall Permeability in Mycobacterium tuberculosis
Article Snippet: Wild-type M. tuberculosis CDC1551 was grown in Middlebrook 7H9 broth (Difco, Sparks, MD) supplemented with 10% oleic acid-albumin-dextrose-catalase (OADC) (Difco), 0.1% glycerol, and 0.05% Tween 80 at 37°C on a roller.

Article Title: Differential expression of iron-, carbon-, and oxygen-responsive mycobacterial genes in the lungs of chronically infected mice and tuberculosis patients
Article Snippet: MTB H37Rv and clinical strains were grown in plastic roller bottles at 37°C in Middlebrook 7H9 broth (Difco) containing 10% oleic acid-albumin-dextrose-catalase (OADC) (Difco), 0.5% glycerol, and 0.05% Tween 80.

Article Title: LprG-Mediated Surface Expression of Lipoarabinomannan Is Essential for Virulence of Mycobacterium tuberculosis
Article Snippet: Bacteriologic media M. tuberculosis was grown in Middlebrook 7H9 broth (DifCo) supplemented with 0.2% glycerol, 10% OADC (DifCo), and 0.05% Tween-80.

Article Title: Rapid Colorimetric Method for Testing Susceptibility of Mycobacterium tuberculosis to Isoniazid and Rifampin in Liquid Cultures
Article Snippet: Several loopfuls of growth from 14-day-old cultures on LJ medium were transferred to sterile tubes with glass beads containing 4 ml of Middlebrook 7H9 broth (Difco, Detroit, Mich.) supplemented with 10% oleic acid-albumin-dextrose-catalase (OADC), 0.5% glycerol, 0.05% Tween 80, and 850 μg of NaNO3 /ml (7H9-S broth).

Infection:

Article Title: Mycobacterium tuberculosis CDC1551 Is Resistant to Reactive Nitrogen and Oxygen Intermediates In Vitro
Article Snippet: Infection with strain CB3.3 was epidemiologically associated with injection drug users with tuberculosis in New York City ( ). .. All strains of mycobacteria were grown in Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) with 0.5% glycerol, 0.02% Tween 80, and ADC enrichment (Difco Laboratories) to mid-log phase.

Article Title: Mimicry of the Pathogenic Mycobacterium Vacuole In Vitro Elicits the Bacterial Intracellular Phenotype, Including Early-Onset Macrophage Death ▿
Article Snippet: .. Five hundred milliliters of Middlebrook 7H9 broth (Difco, Becton-Dickinson, Sparks, MD) was prepared as usual, and then the amounts of the chemicals or supplements listed in were added to make a mixture that mimicked the vacuole 1 h after infection (1-h elemental mixture) and a mixture matching the vacuole 1 day after infection (24-h elemental mixture). .. All chemicals or supplements were from Sigma (St. Louis, MO), except for CaCl2 , which was from Mallinckrodt (St. Louis, MO).

Modification:

Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿
Article Snippet: The M. avium subsp. paratuberculosis strains were prepared for testing as described by Whan et al. , with slight modification. .. Two cryobeads were inoculated into 10 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, MI) supplemented with 0.05% (wt/vol) Tween 80 (Sigma), 10% (vol/vol) Middlebrook oleic acid-albumin-dextrose-catalase supplement (Difco), and 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), pH 6.6 ± 0.2, and the broths were incubated at 37°C with gentle shaking (100 rpm) for 3 to 4 weeks until the optical density at 600 nm (OD600 ) was between 0.7 and 0.9.

Electroporation:

Article Title: Reversible Lipid Accumulation and Associated Division Arrest of Mycobacterium avium in Lipoprotein-Induced Foamy Macrophages May Resemble Key Events during Latency and Reactivation of Tuberculosis
Article Snippet: M. avium TMC 724 was grown at 37°C in Middlebrook 7H9 broth (Difco) supplemented with 0.05% Tween 80 and 10% ADC (Difco). .. Electroporation was performed in cuvettes with 0.2-cm-gap width (Molecular BioProducts [MBP]) at room temperature using a gene pulser (Bio-Rad) with settings of 2.5 kV, 25 μF, and 200 Ω ( ).

Injection:

Article Title: Mycobacterium tuberculosis CDC1551 Is Resistant to Reactive Nitrogen and Oxygen Intermediates In Vitro
Article Snippet: Infection with strain CB3.3 was epidemiologically associated with injection drug users with tuberculosis in New York City ( ). .. All strains of mycobacteria were grown in Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) with 0.5% glycerol, 0.02% Tween 80, and ADC enrichment (Difco Laboratories) to mid-log phase.

Recombinant:

Article Title: LipG a bifunctional phospholipase/thioesterase involved in mycobacterial envelope remodeling
Article Snippet: M. smegmatis mc2 155 and M. smegmatis mc2 155 groEL1ΔC [ ] strains were cultured in Middlebrook 7H9 broth (BD Difco, Le Pont-de-Claix, France) supplemented with 0.05% (v/v ) Tween-80 and 0.2% (v/v ) glycerol (Sigma–Aldrich, Saint-Quentin Fallavier, France) (7H9-S). .. Hygromycin B (Euromedex) was used at a final concentration of 200 and 50 µg/ml for recombinant E. coli and recombinant mycobacteria, respectively.

Transformation Assay:

Article Title: Development of a repressible mycobacterial promoter system based on two transcriptional repressors
Article Snippet: Paragraph title: Bacterial strains, growth media and transformation conditions ... Mycobacterial strains were grown at 37°C in Middlebrook 7H9 broth (Difco) in 150 ml roller bottles with slow rotation (3 rmp) or 7H10 agar plates (Difco), supplemented with 0.2% glycerol and 0.05% Tween-80.

Plasmid Preparation:

Article Title: Reversible Lipid Accumulation and Associated Division Arrest of Mycobacterium avium in Lipoprotein-Induced Foamy Macrophages May Resemble Key Events during Latency and Reactivation of Tuberculosis
Article Snippet: M. avium TMC 724 was grown at 37°C in Middlebrook 7H9 broth (Difco) supplemented with 0.05% Tween 80 and 10% ADC (Difco). .. Three micrograms of plasmid DNA (pCG211) ( ) was added to 200-μl aliquots of electrocompetent bacteria.

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    Difco middlebrook 7h9 broth
    Sources of RNA samples. ( A ) MTB H37Rv was grown in <t>Middlebrook</t> <t>7H9</t> medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.
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    Sources of RNA samples. ( A ) MTB H37Rv was grown in Middlebrook 7H9 medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Differential expression of iron-, carbon-, and oxygen-responsive mycobacterial genes in the lungs of chronically infected mice and tuberculosis patients

    doi: 10.1073/pnas.2436197100

    Figure Lengend Snippet: Sources of RNA samples. ( A ) MTB H37Rv was grown in Middlebrook 7H9 medium. Samples were withdrawn for OD 600 measurements and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, OD 600 ; □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( B ) C57BL/6 mice were aerosol-infected with ≈500 cfu of MTB H37Rv. Lungs were collected for cfu enumeration and RNA extraction; sigA mRNA and 16S rRNA were quantified by QRT-PCR. •, cfu ± SD ( n ≥ 4); □, sigA /16S ratio × 10,000 ± SD ( n ≥ 3). ( C ) Multilayered organization of granulomatous tissue bordering a 4-cm-diameter cavity in the left upper lung lobe of patient 2 (hematoxylin/eosin-stained). At the periphery were residual airspace and lymphocytic aggregates ( i ). A capsule of fibroblasts and fibrin with scattered epithelioid macrophages ( ii ) surrounded a cellular zone containing clusters of multinucleated giant cells ( iii ). Giant cells were also prominent at the border between the cellular zone and the underlying zone of caseation necrosis ( iv ). The surface of the cavity wall bordering on the necrotic zone showed extensive leukocytic infiltration and numerous acid-fast bacilli, not revealed by hematoxylin/eosin staining ( v ). Magnification: ×4( Left ), ×40 ( Right ). Similar pathology was observed in lung specimens from all four patients analyzed in this study.

    Article Snippet: MTB H37Rv and clinical strains were grown in plastic roller bottles at 37°C in Middlebrook 7H9 broth (Difco) containing 10% oleic acid-albumin-dextrose-catalase (OADC) (Difco), 0.5% glycerol, and 0.05% Tween 80.

    Techniques: RNA Extraction, Quantitative RT-PCR, Mouse Assay, Infection, Staining

    (A) Resistance of M. smegmatis to CuOOH is dependent in part upon Ohr. Cells were cultured to late stationary phase (17 days) in Middlebrook 7H9 medium. Cells were then suspended in the same medium at an A 600 of 0.1 and incubated with CuOOH for 2 h at

    Journal: Journal of Bacteriology

    Article Title: Organic Hydroperoxide Resistance Protein and Ergothioneine Compensate for Loss of Mycothiol in Mycobacterium smegmatis Mutants ▿ Mutants ▿ †

    doi: 10.1128/JB.01402-10

    Figure Lengend Snippet: (A) Resistance of M. smegmatis to CuOOH is dependent in part upon Ohr. Cells were cultured to late stationary phase (17 days) in Middlebrook 7H9 medium. Cells were then suspended in the same medium at an A 600 of 0.1 and incubated with CuOOH for 2 h at

    Article Snippet: M. smegmatis mc2 155 was grown in Middlebrook 7H9 broth (Difco) with 0.05% Tween 80 and supplemented with either OADC (oleic acid, albumin, glucose, and catalase supplement) or 1% glucose.

    Techniques: Cell Culture, Incubation

    Clump size distributions in Middlebrook 7H9 broth suspensions of three M. avium subsp. paratuberculosis strains determined using Mastersizer.

    Journal: Applied and Environmental Microbiology

    Article Title: Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk

    doi: 10.1128/AEM.71.6.2853-2861.2005

    Figure Lengend Snippet: Clump size distributions in Middlebrook 7H9 broth suspensions of three M. avium subsp. paratuberculosis strains determined using Mastersizer.

    Article Snippet: Inoculum for the pilot plant runs was prepared at QUB by inoculating 50 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) containing 10% OADC supplement (Difco), 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), and 0.05% Tween 80 (Sigma Chemical Co. Ltd., Poole, England) with a colony of the appropriate strain of M. avium subsp. paratuberculosis from a Herrold's egg yolk medium (HEYM) slope culture and incubating for 8 weeks at 37°C.

    Techniques:

    Clump size distribution in (A) an untreated Middlebrook 7H9 broth suspension of M. avium subsp. paratuberculosis NCTC 8578, (B) the same broth culture homogenized at 2,500 lb/in 2 and analyzed immediately, and (C) the same broth culture analyzed 10 min

    Journal: Applied and Environmental Microbiology

    Article Title: Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk

    doi: 10.1128/AEM.71.6.2853-2861.2005

    Figure Lengend Snippet: Clump size distribution in (A) an untreated Middlebrook 7H9 broth suspension of M. avium subsp. paratuberculosis NCTC 8578, (B) the same broth culture homogenized at 2,500 lb/in 2 and analyzed immediately, and (C) the same broth culture analyzed 10 min

    Article Snippet: Inoculum for the pilot plant runs was prepared at QUB by inoculating 50 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, Mich.) containing 10% OADC supplement (Difco), 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), and 0.05% Tween 80 (Sigma Chemical Co. Ltd., Poole, England) with a colony of the appropriate strain of M. avium subsp. paratuberculosis from a Herrold's egg yolk medium (HEYM) slope culture and incubating for 8 weeks at 37°C.

    Techniques:

    Effects of different concentrations (μg/ml) of trans -cinnamaldehyde and noninhibitory concentrations of ethanol (0.4%) (negative control) on the growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth.

    Journal: Applied and Environmental Microbiology

    Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿

    doi: 10.1128/AEM.00981-08

    Figure Lengend Snippet: Effects of different concentrations (μg/ml) of trans -cinnamaldehyde and noninhibitory concentrations of ethanol (0.4%) (negative control) on the growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth.

    Article Snippet: Two cryobeads were inoculated into 10 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, MI) supplemented with 0.05% (wt/vol) Tween 80 (Sigma), 10% (vol/vol) Middlebrook oleic acid-albumin-dextrose-catalase supplement (Difco), and 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), pH 6.6 ± 0.2, and the broths were incubated at 37°C with gentle shaking (100 rpm) for 3 to 4 weeks until the optical density at 600 nm (OD600 ) was between 0.7 and 0.9.

    Techniques: Negative Control

    Growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth supplemented with the six active compounds at their MICs ( trans -cinnamaldehyde [25.9 μg/ml], cinnamon oil [26.2 μg/ml], carvacrol [72.2 μg/ml], oregano oil [68.2 μg/ml], 2-hydroxy-5-methoxybenzaldehyde [90.4 μg/ml], and 2,5-dihydroxybenzaldehyde [74 μg/ml]), ethanol (0.4%) (negative control), vanillic acid (74 μg/ml), and citral (65.7 μg/ml). 2-Hydroxy-5-methoxybenzaldehyde and 2,5-dihydroxybenzaldehyde had slightly higher initial OD values as they produced colored solutions.

    Journal: Applied and Environmental Microbiology

    Article Title: Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis ▿

    doi: 10.1128/AEM.00981-08

    Figure Lengend Snippet: Growth of M. avium subsp. paratuberculosis NCTC 8578 in Middlebrook 7H9 broth supplemented with the six active compounds at their MICs ( trans -cinnamaldehyde [25.9 μg/ml], cinnamon oil [26.2 μg/ml], carvacrol [72.2 μg/ml], oregano oil [68.2 μg/ml], 2-hydroxy-5-methoxybenzaldehyde [90.4 μg/ml], and 2,5-dihydroxybenzaldehyde [74 μg/ml]), ethanol (0.4%) (negative control), vanillic acid (74 μg/ml), and citral (65.7 μg/ml). 2-Hydroxy-5-methoxybenzaldehyde and 2,5-dihydroxybenzaldehyde had slightly higher initial OD values as they produced colored solutions.

    Article Snippet: Two cryobeads were inoculated into 10 ml Middlebrook 7H9 broth (Difco Laboratories, Detroit, MI) supplemented with 0.05% (wt/vol) Tween 80 (Sigma), 10% (vol/vol) Middlebrook oleic acid-albumin-dextrose-catalase supplement (Difco), and 2 μg/ml mycobactin J (Synbiotics Europe SAS, Lyon, France), pH 6.6 ± 0.2, and the broths were incubated at 37°C with gentle shaking (100 rpm) for 3 to 4 weeks until the optical density at 600 nm (OD600 ) was between 0.7 and 0.9.

    Techniques: Negative Control, Produced