Structured Review

Difco 7h10 solid medium
Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook <t>7H10</t> plates and incubated at 37 °C for 21 days.
7h10 Solid Medium, supplied by Difco, used in various techniques. Bioz Stars score: 91/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Images

1) Product Images from "Assessing the role of Rv1222 (RseA) as an anti-sigma factor of the Mycobacterium tuberculosis extracytoplasmic sigma factor SigE"

Article Title: Assessing the role of Rv1222 (RseA) as an anti-sigma factor of the Mycobacterium tuberculosis extracytoplasmic sigma factor SigE

Journal: Scientific Reports

doi: 10.1038/s41598-019-41183-4

Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook 7H10 plates and incubated at 37 °C for 21 days.
Figure Legend Snippet: Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook 7H10 plates and incubated at 37 °C for 21 days.

Techniques Used: Mutagenesis, Incubation

2) Product Images from "Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source"

Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source

Journal: PLoS Neglected Tropical Diseases

doi: 10.1371/journal.pntd.0002502

Macroscopic phenotypes of M. ulcerans 1615 strain grown on carbohydrates-containing 7H10 media. A- Phenotypes on 7H10 plates and picture of the corresponding pellets. B- Quantification of aggregative abilities of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch by measuring the decrease in OD 600 upon sedimentation of bacterial aggregates in static liquid cultures. Optical density has been measured and compared to initial optical density (t = 0). Relative OD 600 is indicated. C- Electrophoretic mobility of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch, measured according to NaCl concentration in the suspending medium at constant pH.
Figure Legend Snippet: Macroscopic phenotypes of M. ulcerans 1615 strain grown on carbohydrates-containing 7H10 media. A- Phenotypes on 7H10 plates and picture of the corresponding pellets. B- Quantification of aggregative abilities of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch by measuring the decrease in OD 600 upon sedimentation of bacterial aggregates in static liquid cultures. Optical density has been measured and compared to initial optical density (t = 0). Relative OD 600 is indicated. C- Electrophoretic mobility of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch, measured according to NaCl concentration in the suspending medium at constant pH.

Techniques Used: Sedimentation, Concentration Assay

Effect of carbohydrates on mycolactone biosynthesis gene expression in M. ulcerans . A- Schematic representation of mycolactone biosynthesis genes and representation of their function on the mycolactone structure [9] , [10] . The six genes coding for proteins involved in mycolactone synthesis are located on the giant plasmid named pMUM001. The genes mlsA1 (50,973 bp) and mlsA2 (7,233 bp) encode modular type I polyketide synthase (PKS) required for the biosynthesis of the mycolactone core (highlighted in orange). The side chain enzyme is encoded by mlsB gene (42,393 bp) in pink. The mls genes encode 11 different functional domains that are repeated along the gene. Among these modules, the two load modules (LM, in black) are present at 5′end of mlsA1 and mlsB genes and 15 ketoreductase (KR, in grey) are present. There are also three genes coding for potential polyketide-modifying enzymes, including a P450 hydroxylase ( mup053 , in purple), probably responsible for hydroxylation at carbon 12 of the side chain, a potential FabH-like type III ketosynthases (KS) with an acyltransferase activity that might catalyze the C–O bond between the mycolactone core and side-chain ( mup045 , in green) and a putative type II thioesterase ( mup038 , in blue) that may be required for removal of short acyl chains from the PKS loading modules, arising by aberrant decarboxylation. B- Analysis of mycolactone-associated gene expression in M. ulcerans grown on various 7.5% sugar-enriched medium by qRT-PCR. Gene expression has been normalized to the constitutively expressed housekeeping gene ppk encoding the polyphosphate kinase. Relative genes expression is expressed as fold-changes relative to the 7H10 medium. Data are presented as the mean and SD of at least three biological repeats. LM, load module domain.
Figure Legend Snippet: Effect of carbohydrates on mycolactone biosynthesis gene expression in M. ulcerans . A- Schematic representation of mycolactone biosynthesis genes and representation of their function on the mycolactone structure [9] , [10] . The six genes coding for proteins involved in mycolactone synthesis are located on the giant plasmid named pMUM001. The genes mlsA1 (50,973 bp) and mlsA2 (7,233 bp) encode modular type I polyketide synthase (PKS) required for the biosynthesis of the mycolactone core (highlighted in orange). The side chain enzyme is encoded by mlsB gene (42,393 bp) in pink. The mls genes encode 11 different functional domains that are repeated along the gene. Among these modules, the two load modules (LM, in black) are present at 5′end of mlsA1 and mlsB genes and 15 ketoreductase (KR, in grey) are present. There are also three genes coding for potential polyketide-modifying enzymes, including a P450 hydroxylase ( mup053 , in purple), probably responsible for hydroxylation at carbon 12 of the side chain, a potential FabH-like type III ketosynthases (KS) with an acyltransferase activity that might catalyze the C–O bond between the mycolactone core and side-chain ( mup045 , in green) and a putative type II thioesterase ( mup038 , in blue) that may be required for removal of short acyl chains from the PKS loading modules, arising by aberrant decarboxylation. B- Analysis of mycolactone-associated gene expression in M. ulcerans grown on various 7.5% sugar-enriched medium by qRT-PCR. Gene expression has been normalized to the constitutively expressed housekeeping gene ppk encoding the polyphosphate kinase. Relative genes expression is expressed as fold-changes relative to the 7H10 medium. Data are presented as the mean and SD of at least three biological repeats. LM, load module domain.

Techniques Used: Expressing, Plasmid Preparation, Functional Assay, Activity Assay, Quantitative RT-PCR

Total lipids analysis of M. ulcerans . A- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH/H 2 O [20∶4∶0.5, by volume] as the solvent system. Lipids were revealed with alpha-naphthol (left panel) or cupric sulfate (right panel) followed by heating. B- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH [98∶2, by volume] as the solvent system. TLC are shown before (left panel) and after revelation with cupric sulfate and heating (right panel). Arrows highlight new lipid forms in the bacteria grown in 7.5% glucose and maltose-enriched media compared to regular 7H10. Gluc, 7H10 glucose; Malto, 7H10 maltose; MP, 7H10 maltopentaose; starch, 7H10 starch. TDM, trehalose dimycolate; TMM, trehalose monomycolate; GMM, glucose monomycolate; MMM, maltose monomycolate. The compound migrating between TMM and TDM in glucose-grown bacteria does not stain with alpha-naphthol indicating that it is not a glycolipid. Its identity was not determined here.
Figure Legend Snippet: Total lipids analysis of M. ulcerans . A- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH/H 2 O [20∶4∶0.5, by volume] as the solvent system. Lipids were revealed with alpha-naphthol (left panel) or cupric sulfate (right panel) followed by heating. B- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH [98∶2, by volume] as the solvent system. TLC are shown before (left panel) and after revelation with cupric sulfate and heating (right panel). Arrows highlight new lipid forms in the bacteria grown in 7.5% glucose and maltose-enriched media compared to regular 7H10. Gluc, 7H10 glucose; Malto, 7H10 maltose; MP, 7H10 maltopentaose; starch, 7H10 starch. TDM, trehalose dimycolate; TMM, trehalose monomycolate; GMM, glucose monomycolate; MMM, maltose monomycolate. The compound migrating between TMM and TDM in glucose-grown bacteria does not stain with alpha-naphthol indicating that it is not a glycolipid. Its identity was not determined here.

Techniques Used: Thin Layer Chromatography, Staining

Mycolactone quantification by HPLC from M. ulcerans . Relative quantities have been calculated after normalization to the proteins quantities in each sample. The percentages compared to the 7H10 medium are indicated in the chart. *** p
Figure Legend Snippet: Mycolactone quantification by HPLC from M. ulcerans . Relative quantities have been calculated after normalization to the proteins quantities in each sample. The percentages compared to the 7H10 medium are indicated in the chart. *** p

Techniques Used: High Performance Liquid Chromatography

Related Articles

DNA Extraction:

Article Title: Systems Biology-Based Identification of Mycobacterium tuberculosis Persistence Genes in Mouse Lungs
Article Snippet: .. Both lungs were homogenized in phosphate-buffered saline (PBS), plated on supplemented Middlebrook 7H10 solid medium (Difco) containing 20 µg of kanamycin/ml, and incubated at 37°C at least 3 weeks before colony enumeration or DNA extraction. ..

Clone Assay:

Article Title: Genetic Mimetics of Mycobacterium tuberculosis and Methicillin-Resistant Staphylococcus aureus as Verification Standards for Molecular Diagnostics
Article Snippet: Cloning steps were performed in Escherichia coli strain DH5α, and the resulting shuttle plasmids were electroporated into M. smegmatis mc2 155 as previously described ( ). .. M. smegmatis strains were grown at 37°C shaking in Middlebrook 7H9 liquid medium (Difco) supplemented with 0.085% NaCl, 0.2% glucose, 0.2% glycerol, and 0.05% Tween 80 or on Middlebrook 7H10 solid medium (Difco) supplemented with 0.085% NaCl, 0.2% glucose, and 0.5% glycerol.

Article Title: Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Article Snippet: Escherichia coli strain DH5α was used as the host strain for cloning experiments. .. M. smegmatis was also grown on Middlebrook 7H10 solid medium (Difco) supplemented with OADC or 1% glucose.

Centrifugation:

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: After centrifugation (2000 × g for 5 min), the tissue was suspended in PBS with 0.1% Tween 80 and centrifuged again (14000 × g for 10 min). .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days.

In Vitro:

Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source
Article Snippet: Paragraph title: In vitro growth conditions of M. ulcerans 1615 strain ... For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

Mutagenesis:

Article Title: Functional Dissection of the PE Domain Responsible for Translocation of PE_PGRS33 across the Mycobacterial Cell Wall
Article Snippet: M. marinum wild-type strain E11 and its ESX-5 mutant 7C1 were grown at 30°C. .. All mycobacterial strains were grown in Middlebrook 7H9 broth or on 7H10 solid medium (Difco Becton-Dickinson), supplemented with 0.2% glycerol (Sigma-Aldrich), ADC 10% (Becton-Dickinson), and 0.05% v/v Tween 80 (Sigma-Aldrich).

Article Title: Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Article Snippet: M. smegmatis was also grown on Middlebrook 7H10 solid medium (Difco) supplemented with OADC or 1% glucose. .. Complements of the mutant harboring the recombinant pALACE vector were grown on Middlebrook 7H10 solid medium supplemented with OADC and 1% acetamide for induction of the cloned gene.

Isolation:

Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source
Article Snippet: In vitro growth conditions of M. ulcerans 1615 strain The M. ulcerans 1615 strain (Trudeau Collection Strain) originally isolated from human skin biopsies from Malaysia was used to study growth stimulation by various carbohydrates. .. For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

In Vivo:

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: Inoculation and in vivo recombinant vector stability - Groups of seven six-week-old female BALB/c mice were inoculated intraperitoneally with 106 colony forming units (CFU) of BCG Pasteur-85B, BCG Pasteur-85BT, ∆leu D BCG-85B, or ∆leu D BCG-85BT. .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days.

Infection:

Article Title: Role of Cathepsins in Mycobacterium tuberculosis Survival in Human Macrophages
Article Snippet: When required, macrophages were stimulated with 100 IU/ml IFNγ, 18 h prior to infection. .. MTB H37Rv was grown in Middlebrook’s 7H9 medium or 7H10 solid medium and supplemented with 10% OADC Enrichment (Difco) .

Article Title: Differential Transcriptional Response in Macrophages Infected with Cell Wall Deficient versus Normal Mycobacterium Tuberculosis
Article Snippet: Paragraph title: Infection of cell line and determination of bacterial load ... Briefly, cells were lysed by sonication and serial 10-fold dilutions of total cell lysates were plated on either Middlebrook 7H9 semisolid medium (Difco BD, Franklin Lakes, NJ) for CWD-Mtb growth or Middlebrook 7H10 solid medium (Difco BD, Franklin Lakes, NJ) for normal Mtb growth at 37℃.

Article Title: Systems Biology-Based Identification of Mycobacterium tuberculosis Persistence Genes in Mouse Lungs
Article Snippet: Paragraph title: Mouse infection. ... Both lungs were homogenized in phosphate-buffered saline (PBS), plated on supplemented Middlebrook 7H10 solid medium (Difco) containing 20 µg of kanamycin/ml, and incubated at 37°C at least 3 weeks before colony enumeration or DNA extraction.

Mouse Assay:

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: Inoculation and in vivo recombinant vector stability - Groups of seven six-week-old female BALB/c mice were inoculated intraperitoneally with 106 colony forming units (CFU) of BCG Pasteur-85B, BCG Pasteur-85BT, ∆leu D BCG-85B, or ∆leu D BCG-85BT. .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days.

Article Title: Systems Biology-Based Identification of Mycobacterium tuberculosis Persistence Genes in Mouse Lungs
Article Snippet: Five mice per group were sacrificed at days 1, 49, 98, and 196 postinfection. .. Both lungs were homogenized in phosphate-buffered saline (PBS), plated on supplemented Middlebrook 7H10 solid medium (Difco) containing 20 µg of kanamycin/ml, and incubated at 37°C at least 3 weeks before colony enumeration or DNA extraction.

Plasmid Preparation:

Article Title: Role of Cathepsins in Mycobacterium tuberculosis Survival in Human Macrophages
Article Snippet: The strain Mycobacterium smegmatis mc2155, containing a p19 (long lived) EGFP plasmid, was kindly provided by Dr. Douglas Young (London School of Hygiene and Tropical Medicine, London, UK), and the green fluorescent protein (GFP)-expressing strain of M. tuberculosis (H37Rv-pEGFP) plasmid was a kind gift from G. R. Stewart (University of Surrey, United Kingdom). .. MTB H37Rv was grown in Middlebrook’s 7H9 medium or 7H10 solid medium and supplemented with 10% OADC Enrichment (Difco) .

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: Inoculation and in vivo recombinant vector stability - Groups of seven six-week-old female BALB/c mice were inoculated intraperitoneally with 106 colony forming units (CFU) of BCG Pasteur-85B, BCG Pasteur-85BT, ∆leu D BCG-85B, or ∆leu D BCG-85BT. .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days.

Article Title: Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Article Snippet: M. smegmatis was also grown on Middlebrook 7H10 solid medium (Difco) supplemented with OADC or 1% glucose. .. Complements of the mutant harboring the recombinant pALACE vector were grown on Middlebrook 7H10 solid medium supplemented with OADC and 1% acetamide for induction of the cloned gene.

Concentration Assay:

Article Title: Transcriptional Control of the Iron-Responsive fxbA Gene by the Mycobacterial Regulator IdeR †
Article Snippet: Mycobacterial strains were grown on Middlebrook 7H10 solid medium (Difco) and in Middlebrook 7H9 liquid medium (Difco) at 37°C. .. The antibiotics kanamycin and streptomycin, when required, were each added at a concentration of 20 μg/ml.

Incubation:

Article Title: Differential Transcriptional Response in Macrophages Infected with Cell Wall Deficient versus Normal Mycobacterium Tuberculosis
Article Snippet: Briefly, cells were lysed by sonication and serial 10-fold dilutions of total cell lysates were plated on either Middlebrook 7H9 semisolid medium (Difco BD, Franklin Lakes, NJ) for CWD-Mtb growth or Middlebrook 7H10 solid medium (Difco BD, Franklin Lakes, NJ) for normal Mtb growth at 37℃. .. Colonies were counted after either 5 days of incubation for CWD-Mtb growth or 30 days for normal-Mtb growth.

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days. .. The in vivo stability of the constructs was determined by counting the number of colonies on each plate and calculating the percentage of bacteria that retained the recombinant vector, indicated by bacterial growth on the selective medium.

Article Title: Systems Biology-Based Identification of Mycobacterium tuberculosis Persistence Genes in Mouse Lungs
Article Snippet: .. Both lungs were homogenized in phosphate-buffered saline (PBS), plated on supplemented Middlebrook 7H10 solid medium (Difco) containing 20 µg of kanamycin/ml, and incubated at 37°C at least 3 weeks before colony enumeration or DNA extraction. ..

Article Title: Assessing the role of Rv1222 (RseA) as an anti-sigma factor of the Mycobacterium tuberculosis extracytoplasmic sigma factor SigE
Article Snippet: M. tuberculosis H37Rv was grown in either Middlebrook 7H9 liquid medium or Middlebrook 7H10 solid medium (Difco) supplemented with 0.05% Tween 80 and ADN (2% glucose, 5% bovine serum albumin, 0.85% NaCl). .. Plates were incubated at 37 °C in sealed plastic bags.

Construct:

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days. .. The in vivo stability of the constructs was determined by counting the number of colonies on each plate and calculating the percentage of bacteria that retained the recombinant vector, indicated by bacterial growth on the selective medium.

Cell Fractionation:

Article Title: Functional Dissection of the PE Domain Responsible for Translocation of PE_PGRS33 across the Mycobacterial Cell Wall
Article Snippet: All mycobacterial strains were grown in Middlebrook 7H9 broth or on 7H10 solid medium (Difco Becton-Dickinson), supplemented with 0.2% glycerol (Sigma-Aldrich), ADC 10% (Becton-Dickinson), and 0.05% v/v Tween 80 (Sigma-Aldrich). .. Strains processed for the proteinase K assay or cell fractionation were grown in Sauton's medium (Difco) for 14 hours from a starting OD600 of 0.1.

Modification:

Article Title: Cleavage of the moaX-encoded fused molybdopterin synthase from Mycobacterium tuberculosis is necessary for activity
Article Snippet: M. smegmatis strains were grown in Middlebrook 7H9 liquid medium (Difco) supplemented with Middlebrook oleic acid-albumin-dextrose-catalase (OADC) enrichment (Difco), 0.2% glycerol and 0.05% Tween80, with shaking, or on Middlebrook 7H10 solid medium (Difco) supplemented with 0.085% NaCl, 0.2% glucose and 0.5% glycerol. .. Briefly, pre-cultures were washed and inoculated into modified Mycobacterium phlei minimal medium (MPLN), which was modified by excluding asparagine and substituting with 10 mM sodium nitrate, in a final volume of 10 ml to an optical density at 600 nm (OD600 ) of 0.05.

Sonication:

Article Title: Differential Transcriptional Response in Macrophages Infected with Cell Wall Deficient versus Normal Mycobacterium Tuberculosis
Article Snippet: .. Briefly, cells were lysed by sonication and serial 10-fold dilutions of total cell lysates were plated on either Middlebrook 7H9 semisolid medium (Difco BD, Franklin Lakes, NJ) for CWD-Mtb growth or Middlebrook 7H10 solid medium (Difco BD, Franklin Lakes, NJ) for normal Mtb growth at 37℃. .. Colonies were counted after either 5 days of incubation for CWD-Mtb growth or 30 days for normal-Mtb growth.

Recombinant:

Article Title: Stable expression of Mycobacterium bovis antigen 85B in auxotrophic M. bovis bacillus Calmette-Guérin
Article Snippet: Inoculation and in vivo recombinant vector stability - Groups of seven six-week-old female BALB/c mice were inoculated intraperitoneally with 106 colony forming units (CFU) of BCG Pasteur-85B, BCG Pasteur-85BT, ∆leu D BCG-85B, or ∆leu D BCG-85BT. .. The dilutions were plated on 7H10 solid medium supplemented with 10% OADC (Difco) in the presence or absence of selective markers (kanamycin for BCG Pasteur-85B and BCG Pasteur-85BT, and L-leucine for ∆leuD BCG-85B and ∆leuD BCG-85BT) and incubated at 37ºC for 30 days.

Article Title: Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Article Snippet: M. smegmatis was also grown on Middlebrook 7H10 solid medium (Difco) supplemented with OADC or 1% glucose. .. Complements of the mutant harboring the recombinant pALACE vector were grown on Middlebrook 7H10 solid medium supplemented with OADC and 1% acetamide for induction of the cloned gene.

Knock-Out:

Article Title: Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein
Article Snippet: M. smegmatis mc2 155 was the parent wild-type strain used for construction of knockout mutants. .. M. smegmatis was also grown on Middlebrook 7H10 solid medium (Difco) supplemented with OADC or 1% glucose.

Derivative Assay:

Article Title: Rapid Cytolysis of Mycobacterium tuberculosis by Faropenem, an Orally Bioavailable β-Lactam Antibiotic
Article Snippet: .. Mycobacterium tuberculosis Erdman and H37Rv and strains derived from strains Erdman and H37Rv were grown in Middlebrook 7H9 liquid medium (Difco) containing 0.5% albumin, 0.085% NaCl, 0.2% glucose, 0.5% glycerol, and 0.05% Tween 80 or on Middlebrook 7H10 solid medium (Difco) containing 10% oleic acid-albumin-dextrose-catalase (Difco) and 0.5% glycerol. .. Uropathogenic Escherichia coli CFT073 and strains derived from strain CFT073 were grown in LB medium (Sigma).

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  • 93
    Difco middlebrook 7h10 medium
    BCG infection kinetics of murine LN DCs. Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU rBCG-GFP, and LN cells were harvested at different time points. DCs were sorted and analyzed for the presence of rBCG-GFP. Infection of murine LN DCs with BCG ( a ). Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU BCG, and BCG in DCs was quantified in CFUs by culturing on <t>Middlebrook</t> <t>7H10</t> agar ( b ). The results are representative of three independent experiments and presented as means ± SEM. Statistical significance was determined using Student’s t -test (* P
    Middlebrook 7h10 Medium, supplied by Difco, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    7h10  (Difco)
    92
    Difco 7h10
    pimA is an essential gene. Five microliters of a suspension of TB101 was spotted at the indicated dilutions on Middlebrook <t>7H10</t> plates with or without 500 ng/ml ATc.
    7h10, supplied by Difco, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Difco 7h10 solid medium
    Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook <t>7H10</t> plates and incubated at 37 °C for 21 days.
    7h10 Solid Medium, supplied by Difco, used in various techniques. Bioz Stars score: 91/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    BCG infection kinetics of murine LN DCs. Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU rBCG-GFP, and LN cells were harvested at different time points. DCs were sorted and analyzed for the presence of rBCG-GFP. Infection of murine LN DCs with BCG ( a ). Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU BCG, and BCG in DCs was quantified in CFUs by culturing on Middlebrook 7H10 agar ( b ). The results are representative of three independent experiments and presented as means ± SEM. Statistical significance was determined using Student’s t -test (* P

    Journal: BMC Immunology

    Article Title: Rapid loss of early antigen-presenting activity of lymph node dendritic cells against Ag85A protein following Mycobacterium bovis BCG infection

    doi: 10.1186/s12865-018-0258-8

    Figure Lengend Snippet: BCG infection kinetics of murine LN DCs. Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU rBCG-GFP, and LN cells were harvested at different time points. DCs were sorted and analyzed for the presence of rBCG-GFP. Infection of murine LN DCs with BCG ( a ). Six groups of mice ( n = 6) were s.c. injected with 1 × 10 8 CFU BCG, and BCG in DCs was quantified in CFUs by culturing on Middlebrook 7H10 agar ( b ). The results are representative of three independent experiments and presented as means ± SEM. Statistical significance was determined using Student’s t -test (* P

    Article Snippet: Both strains were grown with gentle agitation (80 rpm) in Middlebrook 7H9 medium (Difco, Detroit, MI, USA) supplemented with 0.05% Tween 80 and 10% albumin-dextrose-catalase (ADC) enrichment or on solid Middlebrook 7H10 medium (Difco) supplemented with 0.05% Tween 80 and 10% oleic-ADC enrichment.

    Techniques: Infection, Mouse Assay, Injection

    pimA is an essential gene. Five microliters of a suspension of TB101 was spotted at the indicated dilutions on Middlebrook 7H10 plates with or without 500 ng/ml ATc.

    Journal: Journal of Bacteriology

    Article Title: The Phosphatidyl-myo-Inositol Mannosyltransferase PimA Is Essential for Mycobacterium tuberculosis Growth In Vitro and In Vivo

    doi: 10.1128/JB.01346-13

    Figure Lengend Snippet: pimA is an essential gene. Five microliters of a suspension of TB101 was spotted at the indicated dilutions on Middlebrook 7H10 plates with or without 500 ng/ml ATc.

    Article Snippet: M. tuberculosis H37Rv and its derivative, TB38, were grown at 37°C in Middlebrook 7H9 (liquid medium) or 7H10 (solid medium) (Difco) supplemented with 0.05% (vol/vol) Tween 80 (Sigma-Aldrich), 0.2% (vol/vol) glycerol (Sigma-Aldrich), and 10% ADN (2% glucose, 5% bovine serum albumin, 0.85% NaCl).

    Techniques:

    (A) Growth curves of TB101 in Middlebrook 7H9 containing different concentrations of ATc. The optical density at 540 nm was recorded at different time points and used to compile the growth curves. (B) Killing curves of the pimA conditional mutant TB101 in the presence of ATc. The conditional mutant was grown in Middlebrook 7H9 containing 200 ng/ml ATc. Starting from day 1, samples were collected at different time points and plated on Middlebrook 7H10 to determine the viable counts. Each experiment was repeated at least twice, giving similar results. Filled squares, optical density; open squares, viable counts.

    Journal: Journal of Bacteriology

    Article Title: The Phosphatidyl-myo-Inositol Mannosyltransferase PimA Is Essential for Mycobacterium tuberculosis Growth In Vitro and In Vivo

    doi: 10.1128/JB.01346-13

    Figure Lengend Snippet: (A) Growth curves of TB101 in Middlebrook 7H9 containing different concentrations of ATc. The optical density at 540 nm was recorded at different time points and used to compile the growth curves. (B) Killing curves of the pimA conditional mutant TB101 in the presence of ATc. The conditional mutant was grown in Middlebrook 7H9 containing 200 ng/ml ATc. Starting from day 1, samples were collected at different time points and plated on Middlebrook 7H10 to determine the viable counts. Each experiment was repeated at least twice, giving similar results. Filled squares, optical density; open squares, viable counts.

    Article Snippet: M. tuberculosis H37Rv and its derivative, TB38, were grown at 37°C in Middlebrook 7H9 (liquid medium) or 7H10 (solid medium) (Difco) supplemented with 0.05% (vol/vol) Tween 80 (Sigma-Aldrich), 0.2% (vol/vol) glycerol (Sigma-Aldrich), and 10% ADN (2% glucose, 5% bovine serum albumin, 0.85% NaCl).

    Techniques: Mutagenesis

    Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook 7H10 plates and incubated at 37 °C for 21 days.

    Journal: Scientific Reports

    Article Title: Assessing the role of Rv1222 (RseA) as an anti-sigma factor of the Mycobacterium tuberculosis extracytoplasmic sigma factor SigE

    doi: 10.1038/s41598-019-41183-4

    Figure Lengend Snippet: Growth of M. tuberculosis mutant strains in liquid and solid medium. ( A ) Growth curves in liquid medium. Bacteria were grown in 7H9 ADC at 37 °C in rolling bottles; OD 540 was recorded at regular intervals up to 96 hrs; results represent the average of three independent experiments; ( B ) Growth in solid medium. Bacteria were inoculated in Middlebrook 7H10 plates and incubated at 37 °C for 21 days.

    Article Snippet: M. tuberculosis H37Rv was grown in either Middlebrook 7H9 liquid medium or Middlebrook 7H10 solid medium (Difco) supplemented with 0.05% Tween 80 and ADN (2% glucose, 5% bovine serum albumin, 0.85% NaCl).

    Techniques: Mutagenesis, Incubation

    Macroscopic phenotypes of M. ulcerans 1615 strain grown on carbohydrates-containing 7H10 media. A- Phenotypes on 7H10 plates and picture of the corresponding pellets. B- Quantification of aggregative abilities of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch by measuring the decrease in OD 600 upon sedimentation of bacterial aggregates in static liquid cultures. Optical density has been measured and compared to initial optical density (t = 0). Relative OD 600 is indicated. C- Electrophoretic mobility of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch, measured according to NaCl concentration in the suspending medium at constant pH.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source

    doi: 10.1371/journal.pntd.0002502

    Figure Lengend Snippet: Macroscopic phenotypes of M. ulcerans 1615 strain grown on carbohydrates-containing 7H10 media. A- Phenotypes on 7H10 plates and picture of the corresponding pellets. B- Quantification of aggregative abilities of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch by measuring the decrease in OD 600 upon sedimentation of bacterial aggregates in static liquid cultures. Optical density has been measured and compared to initial optical density (t = 0). Relative OD 600 is indicated. C- Electrophoretic mobility of M. ulcerans 1615 strain grown on 7H10 medium containing 7.5% of glucose or starch, measured according to NaCl concentration in the suspending medium at constant pH.

    Article Snippet: For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

    Techniques: Sedimentation, Concentration Assay

    Effect of carbohydrates on mycolactone biosynthesis gene expression in M. ulcerans . A- Schematic representation of mycolactone biosynthesis genes and representation of their function on the mycolactone structure [9] , [10] . The six genes coding for proteins involved in mycolactone synthesis are located on the giant plasmid named pMUM001. The genes mlsA1 (50,973 bp) and mlsA2 (7,233 bp) encode modular type I polyketide synthase (PKS) required for the biosynthesis of the mycolactone core (highlighted in orange). The side chain enzyme is encoded by mlsB gene (42,393 bp) in pink. The mls genes encode 11 different functional domains that are repeated along the gene. Among these modules, the two load modules (LM, in black) are present at 5′end of mlsA1 and mlsB genes and 15 ketoreductase (KR, in grey) are present. There are also three genes coding for potential polyketide-modifying enzymes, including a P450 hydroxylase ( mup053 , in purple), probably responsible for hydroxylation at carbon 12 of the side chain, a potential FabH-like type III ketosynthases (KS) with an acyltransferase activity that might catalyze the C–O bond between the mycolactone core and side-chain ( mup045 , in green) and a putative type II thioesterase ( mup038 , in blue) that may be required for removal of short acyl chains from the PKS loading modules, arising by aberrant decarboxylation. B- Analysis of mycolactone-associated gene expression in M. ulcerans grown on various 7.5% sugar-enriched medium by qRT-PCR. Gene expression has been normalized to the constitutively expressed housekeeping gene ppk encoding the polyphosphate kinase. Relative genes expression is expressed as fold-changes relative to the 7H10 medium. Data are presented as the mean and SD of at least three biological repeats. LM, load module domain.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source

    doi: 10.1371/journal.pntd.0002502

    Figure Lengend Snippet: Effect of carbohydrates on mycolactone biosynthesis gene expression in M. ulcerans . A- Schematic representation of mycolactone biosynthesis genes and representation of their function on the mycolactone structure [9] , [10] . The six genes coding for proteins involved in mycolactone synthesis are located on the giant plasmid named pMUM001. The genes mlsA1 (50,973 bp) and mlsA2 (7,233 bp) encode modular type I polyketide synthase (PKS) required for the biosynthesis of the mycolactone core (highlighted in orange). The side chain enzyme is encoded by mlsB gene (42,393 bp) in pink. The mls genes encode 11 different functional domains that are repeated along the gene. Among these modules, the two load modules (LM, in black) are present at 5′end of mlsA1 and mlsB genes and 15 ketoreductase (KR, in grey) are present. There are also three genes coding for potential polyketide-modifying enzymes, including a P450 hydroxylase ( mup053 , in purple), probably responsible for hydroxylation at carbon 12 of the side chain, a potential FabH-like type III ketosynthases (KS) with an acyltransferase activity that might catalyze the C–O bond between the mycolactone core and side-chain ( mup045 , in green) and a putative type II thioesterase ( mup038 , in blue) that may be required for removal of short acyl chains from the PKS loading modules, arising by aberrant decarboxylation. B- Analysis of mycolactone-associated gene expression in M. ulcerans grown on various 7.5% sugar-enriched medium by qRT-PCR. Gene expression has been normalized to the constitutively expressed housekeeping gene ppk encoding the polyphosphate kinase. Relative genes expression is expressed as fold-changes relative to the 7H10 medium. Data are presented as the mean and SD of at least three biological repeats. LM, load module domain.

    Article Snippet: For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

    Techniques: Expressing, Plasmid Preparation, Functional Assay, Activity Assay, Quantitative RT-PCR

    Total lipids analysis of M. ulcerans . A- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH/H 2 O [20∶4∶0.5, by volume] as the solvent system. Lipids were revealed with alpha-naphthol (left panel) or cupric sulfate (right panel) followed by heating. B- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH [98∶2, by volume] as the solvent system. TLC are shown before (left panel) and after revelation with cupric sulfate and heating (right panel). Arrows highlight new lipid forms in the bacteria grown in 7.5% glucose and maltose-enriched media compared to regular 7H10. Gluc, 7H10 glucose; Malto, 7H10 maltose; MP, 7H10 maltopentaose; starch, 7H10 starch. TDM, trehalose dimycolate; TMM, trehalose monomycolate; GMM, glucose monomycolate; MMM, maltose monomycolate. The compound migrating between TMM and TDM in glucose-grown bacteria does not stain with alpha-naphthol indicating that it is not a glycolipid. Its identity was not determined here.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source

    doi: 10.1371/journal.pntd.0002502

    Figure Lengend Snippet: Total lipids analysis of M. ulcerans . A- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH/H 2 O [20∶4∶0.5, by volume] as the solvent system. Lipids were revealed with alpha-naphthol (left panel) or cupric sulfate (right panel) followed by heating. B- Lipids from M. ulcerans were separated by TLC using CHCl 3 /CH 3 OH [98∶2, by volume] as the solvent system. TLC are shown before (left panel) and after revelation with cupric sulfate and heating (right panel). Arrows highlight new lipid forms in the bacteria grown in 7.5% glucose and maltose-enriched media compared to regular 7H10. Gluc, 7H10 glucose; Malto, 7H10 maltose; MP, 7H10 maltopentaose; starch, 7H10 starch. TDM, trehalose dimycolate; TMM, trehalose monomycolate; GMM, glucose monomycolate; MMM, maltose monomycolate. The compound migrating between TMM and TDM in glucose-grown bacteria does not stain with alpha-naphthol indicating that it is not a glycolipid. Its identity was not determined here.

    Article Snippet: For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

    Techniques: Thin Layer Chromatography, Staining

    Mycolactone quantification by HPLC from M. ulcerans . Relative quantities have been calculated after normalization to the proteins quantities in each sample. The percentages compared to the 7H10 medium are indicated in the chart. *** p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Regulation of Mycolactone, the Mycobacterium ulcerans Toxin, Depends on Nutrient Source

    doi: 10.1371/journal.pntd.0002502

    Figure Lengend Snippet: Mycolactone quantification by HPLC from M. ulcerans . Relative quantities have been calculated after normalization to the proteins quantities in each sample. The percentages compared to the 7H10 medium are indicated in the chart. *** p

    Article Snippet: For culture on solid medium, M. ulcerans 1615 strain was cultivated onto 7H10 solid medium supplemented with 10% OADC (oleic acid, dextrose, catalase; Difco, Becton-Dickinson) at 30°C for one month.

    Techniques: High Performance Liquid Chromatography