hot firepol evagreen hrm mix  (Solis BioDyne)


Bioz Verified Symbol Solis BioDyne is a verified supplier
Bioz Manufacturer Symbol Solis BioDyne manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 96

    Structured Review

    Solis BioDyne hot firepol evagreen hrm mix
    Blood-meal <t>HRM</t> profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and <t>mixed</t> species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.
    Hot Firepol Evagreen Hrm Mix, supplied by Solis BioDyne, used in various techniques. Bioz Stars score: 96/100, based on 40 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hot firepol evagreen hrm mix/product/Solis BioDyne
    Average 96 stars, based on 40 article reviews
    Price from $9.99 to $1999.99
    hot firepol evagreen hrm mix - by Bioz Stars, 2022-08
    96/100 stars

    Images

    1) Product Images from "Tsetse blood-meal sources, endosymbionts and trypanosome-associations in the Maasai Mara National Reserve, a wildlife-human-livestock interface"

    Article Title: Tsetse blood-meal sources, endosymbionts and trypanosome-associations in the Maasai Mara National Reserve, a wildlife-human-livestock interface

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0008267

    Blood-meal HRM profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.
    Figure Legend Snippet: Blood-meal HRM profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.

    Techniques Used:

    2) Product Images from "Detection of Species Substitution in the Meat Value Chain by High-Resolution Melting Analysis of Mitochondrial PCR Products"

    Article Title: Detection of Species Substitution in the Meat Value Chain by High-Resolution Melting Analysis of Mitochondrial PCR Products

    Journal: Foods

    doi: 10.3390/foods10123090

    PCR-HRM melt rate profiles of pure and mixed meat samples assessed using three mitochondrial markers. The left column represents red meat sources (sheep, goat, cattle, and camel) and their corresponding mixtures, whereas the right column represents DNA from white meat sources (Nile perch, chicken, and pig) and their mixtures. Distinct melt rates are represented as changes in fluorescence units with increasing temperatures (dF/dT) for ( a ) CO1 , ( b ) cyt b , and ( c ) 16S rRNA markers.
    Figure Legend Snippet: PCR-HRM melt rate profiles of pure and mixed meat samples assessed using three mitochondrial markers. The left column represents red meat sources (sheep, goat, cattle, and camel) and their corresponding mixtures, whereas the right column represents DNA from white meat sources (Nile perch, chicken, and pig) and their mixtures. Distinct melt rates are represented as changes in fluorescence units with increasing temperatures (dF/dT) for ( a ) CO1 , ( b ) cyt b , and ( c ) 16S rRNA markers.

    Techniques Used: Polymerase Chain Reaction, Fluorescence

    3) Product Images from "Tsetse blood-meal sources, endosymbionts, and trypanosome infections provide insight into African trypanosomiasis transmission in the Maasai Mara National Reserve, a wildlife-human-livestock interface"

    Article Title: Tsetse blood-meal sources, endosymbionts, and trypanosome infections provide insight into African trypanosomiasis transmission in the Maasai Mara National Reserve, a wildlife-human-livestock interface

    Journal: bioRxiv

    doi: 10.1101/2020.04.06.027367

    Blood-meal melt curves and proportions of vertebrate species identified. A. High resolution melt curves of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt profile peaks to those of more than one blood-meal control. B. Overall blood-meal proportions and proportions per tsetse species.
    Figure Legend Snippet: Blood-meal melt curves and proportions of vertebrate species identified. A. High resolution melt curves of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt profile peaks to those of more than one blood-meal control. B. Overall blood-meal proportions and proportions per tsetse species.

    Techniques Used:

    4) Product Images from "Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer"

    Article Title: Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer

    Journal: BMC Cancer

    doi: 10.1186/1471-2407-13-526

    DNA methylation and expression level of the PHD3 gene in HCT116 and DLD-1 CRC cells. A. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles. B. Cells were cultured in DMEM either in hypoxic (1%O 2 ) or normoxic conditions for 48 hrs. After incubation, the cells were used for total RNA isolation and reverse transcription. The PHD3 cDNA levels were determined by RQ-PCR relative quantification analysis. RQ-PCR results were standardized by the geometric mean of PBGD and hMRPL19 cDNA levels. PHD3 cDNA levels are expressed as a multiplicity of these cDNA copies in the cell line’s calibrator. C. Cells were cultured in DMEM either in hypoxic (1%O 2 ) ( H ) or normoxic ( N ) conditions for 48 hrs. Cells were then used for protein isolation. Proteins were separated by 10% SDS-PAGE, and transferred to a membrane that was then immunoblotted with Rp anti - PHD3 Ab and incubated with goat anti-rabbit HRP-conjugated Ab. The membrane was then stripped and reblotted with Rp anti-GAPDH Ab, followed by incubation with goat anti-rabbit HRP-conjugated Ab. The band densitometry readings were normalized to GAPDH loading control. The ratio of PHD3 to GAPDH for DLD-1 in normoxic conditions was assumed to be 1.
    Figure Legend Snippet: DNA methylation and expression level of the PHD3 gene in HCT116 and DLD-1 CRC cells. A. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles. B. Cells were cultured in DMEM either in hypoxic (1%O 2 ) or normoxic conditions for 48 hrs. After incubation, the cells were used for total RNA isolation and reverse transcription. The PHD3 cDNA levels were determined by RQ-PCR relative quantification analysis. RQ-PCR results were standardized by the geometric mean of PBGD and hMRPL19 cDNA levels. PHD3 cDNA levels are expressed as a multiplicity of these cDNA copies in the cell line’s calibrator. C. Cells were cultured in DMEM either in hypoxic (1%O 2 ) ( H ) or normoxic ( N ) conditions for 48 hrs. Cells were then used for protein isolation. Proteins were separated by 10% SDS-PAGE, and transferred to a membrane that was then immunoblotted with Rp anti - PHD3 Ab and incubated with goat anti-rabbit HRP-conjugated Ab. The membrane was then stripped and reblotted with Rp anti-GAPDH Ab, followed by incubation with goat anti-rabbit HRP-conjugated Ab. The band densitometry readings were normalized to GAPDH loading control. The ratio of PHD3 to GAPDH for DLD-1 in normoxic conditions was assumed to be 1.

    Techniques Used: DNA Methylation Assay, Expressing, Cell Culture, DNA Extraction, Modification, Methylation, Real-time Polymerase Chain Reaction, Amplification, Incubation, Isolation, Polymerase Chain Reaction, SDS Page

    DNA methylation assessment of PHD3 gene regulatory region by bisulfite sequencing and HRM analysis in primary tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC (P1-P5) were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. The PHD3 regions containing 60 CpG dinucleotides (chr14: 34 419 929-34 420 563) (Top panel A ) and 44 CpG dinucleotides (ch14: 34 419 346-34 419 943) (Top panel B ) were then amplified by a pair of primers complementary to the bisulfite-DNA modified sequence (Additional file 1 , Additional file 2 ). The PCR products were purified with subsequent cloning into a plasmid vector. Plasmid DNA isolated from five positive bacterial clones was used for commercial sequencing. The results of bisulfite sequencing were assessed and presented using BiQ analyzer software and BDPC web server [ 23 , 24 ]. Black and grey boxes represent methylated and unmethylated CpG dinucleotide, respectively. Red rectangles correspond to regions amplified in HRM analysis by specific primers PHD3.1 (chr14: 34 419 922-34 420 080), PHD3.2 (chr14: 34 419 795- 34 419 935) and PHD3.3 (chr14: 34 419 400-34 419 538) (Additional file 1 , Additional file 2 ). Bottom panels A and B represent HRM profiles of standard and example of patient DNA (patient P2 from bisulfite sequencing) PCR product. Methylation percentage of three DNA fragments within the PHD3 CpG island was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite treated DNA. HRM methylation analysis was performed using Light Cycler®480 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate.
    Figure Legend Snippet: DNA methylation assessment of PHD3 gene regulatory region by bisulfite sequencing and HRM analysis in primary tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC (P1-P5) were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. The PHD3 regions containing 60 CpG dinucleotides (chr14: 34 419 929-34 420 563) (Top panel A ) and 44 CpG dinucleotides (ch14: 34 419 346-34 419 943) (Top panel B ) were then amplified by a pair of primers complementary to the bisulfite-DNA modified sequence (Additional file 1 , Additional file 2 ). The PCR products were purified with subsequent cloning into a plasmid vector. Plasmid DNA isolated from five positive bacterial clones was used for commercial sequencing. The results of bisulfite sequencing were assessed and presented using BiQ analyzer software and BDPC web server [ 23 , 24 ]. Black and grey boxes represent methylated and unmethylated CpG dinucleotide, respectively. Red rectangles correspond to regions amplified in HRM analysis by specific primers PHD3.1 (chr14: 34 419 922-34 420 080), PHD3.2 (chr14: 34 419 795- 34 419 935) and PHD3.3 (chr14: 34 419 400-34 419 538) (Additional file 1 , Additional file 2 ). Bottom panels A and B represent HRM profiles of standard and example of patient DNA (patient P2 from bisulfite sequencing) PCR product. Methylation percentage of three DNA fragments within the PHD3 CpG island was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite treated DNA. HRM methylation analysis was performed using Light Cycler®480 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate.

    Techniques Used: DNA Methylation Assay, Methylation Sequencing, DNA Extraction, Amplification, Modification, Sequencing, Polymerase Chain Reaction, Purification, Clone Assay, Plasmid Preparation, Isolation, Software, Methylation, Real-time Polymerase Chain Reaction

    Ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level in three ranges of PHD3 methylation status: 0–1%; 1–10% and 10–100%. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. The methylation for each patient was calculated as an average percentage of methylation in amplified fragments located in the CpG island of PHD3. The samples were divided into three groups for statistical analysis: 0–1% methylation, 1–10% methylation and 10–100% methylation (Table 2 ) [ 28 - 30 ]. To evaluate the statistically significant difference in the ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level between the three DNA methylation ranges (0–1% methylation, 1–10% methylation and 10–100% methylation), the non-parametric Kruskal-Wallis test was employed.
    Figure Legend Snippet: Ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level in three ranges of PHD3 methylation status: 0–1%; 1–10% and 10–100%. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. The methylation for each patient was calculated as an average percentage of methylation in amplified fragments located in the CpG island of PHD3. The samples were divided into three groups for statistical analysis: 0–1% methylation, 1–10% methylation and 10–100% methylation (Table 2 ) [ 28 - 30 ]. To evaluate the statistically significant difference in the ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level between the three DNA methylation ranges (0–1% methylation, 1–10% methylation and 10–100% methylation), the non-parametric Kruskal-Wallis test was employed.

    Techniques Used: Methylation, Real-time Polymerase Chain Reaction, Amplification, DNA Methylation Assay

    Effect of 5-dAzaC on PHD3 gene DNA methylation in HCT116 and DLD-1 CRC cells. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions either in the absence or in the presence of 5-dAzaC at a concentration of 5.00 μM for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island: A (chr14: 34 419 922–34 420 080), B (chr14: 34 419 795–34 419 935) and C (chr14: 34 419 400–34 419 538) (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles.
    Figure Legend Snippet: Effect of 5-dAzaC on PHD3 gene DNA methylation in HCT116 and DLD-1 CRC cells. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions either in the absence or in the presence of 5-dAzaC at a concentration of 5.00 μM for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island: A (chr14: 34 419 922–34 420 080), B (chr14: 34 419 795–34 419 935) and C (chr14: 34 419 400–34 419 538) (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles.

    Techniques Used: DNA Methylation Assay, Cell Culture, Concentration Assay, DNA Extraction, Modification, Methylation, Real-time Polymerase Chain Reaction, Amplification

    5) Product Images from "COXIBs and 2,5-dimethylcelecoxib counteract the hyperactivated Wnt/β-catenin pathway and COX-2/PGE2/EP4 signaling in glioblastoma cells"

    Article Title: COXIBs and 2,5-dimethylcelecoxib counteract the hyperactivated Wnt/β-catenin pathway and COX-2/PGE2/EP4 signaling in glioblastoma cells

    Journal: BMC Cancer

    doi: 10.1186/s12885-021-08164-1

    MGMT methylation level in DNA isolated from T98G cells determined using MS-HRM analysis after 48 h of treatment with the analyzed COXIBs, 2,5-DMC, PKF118–310 and 0.5% DMSO (control). The concentration of compounds used in this assay provided at least 70% of cell viability. Normalized melting curves of standardized solutions with a given DNA methylation percentage (indicated with ‘M’) in a background of unmethylated DNA are presented together with the results of the analysis. For clarity, one replicate of each analyzed sample/control is shown
    Figure Legend Snippet: MGMT methylation level in DNA isolated from T98G cells determined using MS-HRM analysis after 48 h of treatment with the analyzed COXIBs, 2,5-DMC, PKF118–310 and 0.5% DMSO (control). The concentration of compounds used in this assay provided at least 70% of cell viability. Normalized melting curves of standardized solutions with a given DNA methylation percentage (indicated with ‘M’) in a background of unmethylated DNA are presented together with the results of the analysis. For clarity, one replicate of each analyzed sample/control is shown

    Techniques Used: Methylation, Isolation, Concentration Assay, DNA Methylation Assay

    6) Product Images from "Tsetse Bloodmeal Analyses Incriminate the Common Warthog Phacochoerus africanus as an Important Cryptic Host of Animal Trypanosomes in Smallholder Cattle Farming Communities in Shimba Hills, Kenya"

    Article Title: Tsetse Bloodmeal Analyses Incriminate the Common Warthog Phacochoerus africanus as an Important Cryptic Host of Animal Trypanosomes in Smallholder Cattle Farming Communities in Shimba Hills, Kenya

    Journal: Pathogens

    doi: 10.3390/pathogens10111501

    (A – E)  High-Resolution Melt profiles of vertebrate bloodmeals in tsetse flies. Profiles are distinguished using different colours to denote different vertebrate bloodmeal hosts. The identity of a vertebrate bloodmeal is shown on the right side of each graph.
    Figure Legend Snippet: (A – E) High-Resolution Melt profiles of vertebrate bloodmeals in tsetse flies. Profiles are distinguished using different colours to denote different vertebrate bloodmeal hosts. The identity of a vertebrate bloodmeal is shown on the right side of each graph.

    Techniques Used:

    7) Product Images from "Clinical significance of DNA methylation mRNA levels of TET family members in colorectal cancer"

    Article Title: Clinical significance of DNA methylation mRNA levels of TET family members in colorectal cancer

    Journal: Journal of Cancer Research and Clinical Oncology

    doi: 10.1007/s00432-014-1901-2

    DNA methylation assessment of TET1 , TET2 and TET3 gene regulatory region by HRM analysis in tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. a – c Represent HRM profiles of standard and example of patient DNA PCR product for TET1, TET2 and TET3, respectively. Methylation percentage of DNA fragments within the CpG island was determined by real-time PCR amplification of bisulfite-treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite-treated standard DNA. HRM methylation analysis was performed using Light Cycler ® 480 or LightCycler ® 96 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate
    Figure Legend Snippet: DNA methylation assessment of TET1 , TET2 and TET3 gene regulatory region by HRM analysis in tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. a – c Represent HRM profiles of standard and example of patient DNA PCR product for TET1, TET2 and TET3, respectively. Methylation percentage of DNA fragments within the CpG island was determined by real-time PCR amplification of bisulfite-treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite-treated standard DNA. HRM methylation analysis was performed using Light Cycler ® 480 or LightCycler ® 96 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate

    Techniques Used: DNA Methylation Assay, DNA Extraction, Polymerase Chain Reaction, Methylation, Real-time Polymerase Chain Reaction, Amplification, Software

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 96
    Solis BioDyne hot firepol evagreen hrm mix
    Blood-meal <t>HRM</t> profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and <t>mixed</t> species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.
    Hot Firepol Evagreen Hrm Mix, supplied by Solis BioDyne, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hot firepol evagreen hrm mix/product/Solis BioDyne
    Average 96 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hot firepol evagreen hrm mix - by Bioz Stars, 2022-08
    96/100 stars
      Buy from Supplier

    Image Search Results


    Blood-meal HRM profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Tsetse blood-meal sources, endosymbionts and trypanosome-associations in the Maasai Mara National Reserve, a wildlife-human-livestock interface

    doi: 10.1371/journal.pntd.0008267

    Figure Lengend Snippet: Blood-meal HRM profiles and proportions of vertebrate species identified. Panel A. HRM profiles of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt rate profiles to those of more than one blood-meal control. Panel B. Overall and per-tsetse-species proportions of vertebrate blood-meal sources.

    Article Snippet: The PCRs were carried out in 20-μl reaction volumes, which included 4 ul of 5× Hot FIREPol EvaGreen HRM Mix (Solis BioDyne, Tartu, Estonia), 0.5 μM of each primer, 50 ng of DNA template, and 10 μl of PCR grade water.

    Techniques:

    PCR-HRM melt rate profiles of pure and mixed meat samples assessed using three mitochondrial markers. The left column represents red meat sources (sheep, goat, cattle, and camel) and their corresponding mixtures, whereas the right column represents DNA from white meat sources (Nile perch, chicken, and pig) and their mixtures. Distinct melt rates are represented as changes in fluorescence units with increasing temperatures (dF/dT) for ( a ) CO1 , ( b ) cyt b , and ( c ) 16S rRNA markers.

    Journal: Foods

    Article Title: Detection of Species Substitution in the Meat Value Chain by High-Resolution Melting Analysis of Mitochondrial PCR Products

    doi: 10.3390/foods10123090

    Figure Lengend Snippet: PCR-HRM melt rate profiles of pure and mixed meat samples assessed using three mitochondrial markers. The left column represents red meat sources (sheep, goat, cattle, and camel) and their corresponding mixtures, whereas the right column represents DNA from white meat sources (Nile perch, chicken, and pig) and their mixtures. Distinct melt rates are represented as changes in fluorescence units with increasing temperatures (dF/dT) for ( a ) CO1 , ( b ) cyt b , and ( c ) 16S rRNA markers.

    Article Snippet: Briefly, 10 µL PCR reactions were set up, each comprising 1× HOT FIREPol® EvaGreen® HRM Mix no ROX (Solis BioDyne, Tartu, Estonia), 0.5 µM of both forward and reverse primers , 20 ng of DNA template, and nuclease-free water.

    Techniques: Polymerase Chain Reaction, Fluorescence

    Blood-meal melt curves and proportions of vertebrate species identified. A. High resolution melt curves of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt profile peaks to those of more than one blood-meal control. B. Overall blood-meal proportions and proportions per tsetse species.

    Journal: bioRxiv

    Article Title: Tsetse blood-meal sources, endosymbionts, and trypanosome infections provide insight into African trypanosomiasis transmission in the Maasai Mara National Reserve, a wildlife-human-livestock interface

    doi: 10.1101/2020.04.06.027367

    Figure Lengend Snippet: Blood-meal melt curves and proportions of vertebrate species identified. A. High resolution melt curves of single species and mixed species blood-meals. Mixed blood-meals were determined by matching melt profile peaks to those of more than one blood-meal control. B. Overall blood-meal proportions and proportions per tsetse species.

    Article Snippet: Final concentrations of the PCR reactions were performed in 20 µl PCR reaction volumes, which included 4 ul of 5× Hot FIREPol EvaGreen HRM Mix (Solis BioDyne, Teaduspargi, Estonia), 0.5 µM of each primer, 250 ng of DNA template, and 10 µl of PCR grade water.

    Techniques:

    DNA methylation and expression level of the PHD3 gene in HCT116 and DLD-1 CRC cells. A. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles. B. Cells were cultured in DMEM either in hypoxic (1%O 2 ) or normoxic conditions for 48 hrs. After incubation, the cells were used for total RNA isolation and reverse transcription. The PHD3 cDNA levels were determined by RQ-PCR relative quantification analysis. RQ-PCR results were standardized by the geometric mean of PBGD and hMRPL19 cDNA levels. PHD3 cDNA levels are expressed as a multiplicity of these cDNA copies in the cell line’s calibrator. C. Cells were cultured in DMEM either in hypoxic (1%O 2 ) ( H ) or normoxic ( N ) conditions for 48 hrs. Cells were then used for protein isolation. Proteins were separated by 10% SDS-PAGE, and transferred to a membrane that was then immunoblotted with Rp anti - PHD3 Ab and incubated with goat anti-rabbit HRP-conjugated Ab. The membrane was then stripped and reblotted with Rp anti-GAPDH Ab, followed by incubation with goat anti-rabbit HRP-conjugated Ab. The band densitometry readings were normalized to GAPDH loading control. The ratio of PHD3 to GAPDH for DLD-1 in normoxic conditions was assumed to be 1.

    Journal: BMC Cancer

    Article Title: Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer

    doi: 10.1186/1471-2407-13-526

    Figure Lengend Snippet: DNA methylation and expression level of the PHD3 gene in HCT116 and DLD-1 CRC cells. A. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles. B. Cells were cultured in DMEM either in hypoxic (1%O 2 ) or normoxic conditions for 48 hrs. After incubation, the cells were used for total RNA isolation and reverse transcription. The PHD3 cDNA levels were determined by RQ-PCR relative quantification analysis. RQ-PCR results were standardized by the geometric mean of PBGD and hMRPL19 cDNA levels. PHD3 cDNA levels are expressed as a multiplicity of these cDNA copies in the cell line’s calibrator. C. Cells were cultured in DMEM either in hypoxic (1%O 2 ) ( H ) or normoxic ( N ) conditions for 48 hrs. Cells were then used for protein isolation. Proteins were separated by 10% SDS-PAGE, and transferred to a membrane that was then immunoblotted with Rp anti - PHD3 Ab and incubated with goat anti-rabbit HRP-conjugated Ab. The membrane was then stripped and reblotted with Rp anti-GAPDH Ab, followed by incubation with goat anti-rabbit HRP-conjugated Ab. The band densitometry readings were normalized to GAPDH loading control. The ratio of PHD3 to GAPDH for DLD-1 in normoxic conditions was assumed to be 1.

    Article Snippet: For PCR amplification, 1 μl of the bisulfite treated DNA from patients, HCT116, DLD-1 cells, or standards, and primers (Additional file , Additional file ) was added to 19 μl of 5 X Hot FIREPol EvaGreen HRM Mix, Solis BioDyne Co. (Tartu, Estonia).

    Techniques: DNA Methylation Assay, Expressing, Cell Culture, DNA Extraction, Modification, Methylation, Real-time Polymerase Chain Reaction, Amplification, Incubation, Isolation, Polymerase Chain Reaction, SDS Page

    DNA methylation assessment of PHD3 gene regulatory region by bisulfite sequencing and HRM analysis in primary tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC (P1-P5) were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. The PHD3 regions containing 60 CpG dinucleotides (chr14: 34 419 929-34 420 563) (Top panel A ) and 44 CpG dinucleotides (ch14: 34 419 346-34 419 943) (Top panel B ) were then amplified by a pair of primers complementary to the bisulfite-DNA modified sequence (Additional file 1 , Additional file 2 ). The PCR products were purified with subsequent cloning into a plasmid vector. Plasmid DNA isolated from five positive bacterial clones was used for commercial sequencing. The results of bisulfite sequencing were assessed and presented using BiQ analyzer software and BDPC web server [ 23 , 24 ]. Black and grey boxes represent methylated and unmethylated CpG dinucleotide, respectively. Red rectangles correspond to regions amplified in HRM analysis by specific primers PHD3.1 (chr14: 34 419 922-34 420 080), PHD3.2 (chr14: 34 419 795- 34 419 935) and PHD3.3 (chr14: 34 419 400-34 419 538) (Additional file 1 , Additional file 2 ). Bottom panels A and B represent HRM profiles of standard and example of patient DNA (patient P2 from bisulfite sequencing) PCR product. Methylation percentage of three DNA fragments within the PHD3 CpG island was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite treated DNA. HRM methylation analysis was performed using Light Cycler®480 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate.

    Journal: BMC Cancer

    Article Title: Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer

    doi: 10.1186/1471-2407-13-526

    Figure Lengend Snippet: DNA methylation assessment of PHD3 gene regulatory region by bisulfite sequencing and HRM analysis in primary tissue samples from patients with CRC. Primary cancerous and histopathologically unchanged tissues from the same patients with CRC (P1-P5) were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. The PHD3 regions containing 60 CpG dinucleotides (chr14: 34 419 929-34 420 563) (Top panel A ) and 44 CpG dinucleotides (ch14: 34 419 346-34 419 943) (Top panel B ) were then amplified by a pair of primers complementary to the bisulfite-DNA modified sequence (Additional file 1 , Additional file 2 ). The PCR products were purified with subsequent cloning into a plasmid vector. Plasmid DNA isolated from five positive bacterial clones was used for commercial sequencing. The results of bisulfite sequencing were assessed and presented using BiQ analyzer software and BDPC web server [ 23 , 24 ]. Black and grey boxes represent methylated and unmethylated CpG dinucleotide, respectively. Red rectangles correspond to regions amplified in HRM analysis by specific primers PHD3.1 (chr14: 34 419 922-34 420 080), PHD3.2 (chr14: 34 419 795- 34 419 935) and PHD3.3 (chr14: 34 419 400-34 419 538) (Additional file 1 , Additional file 2 ). Bottom panels A and B represent HRM profiles of standard and example of patient DNA (patient P2 from bisulfite sequencing) PCR product. Methylation percentage of three DNA fragments within the PHD3 CpG island was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. DNA standards were prepared by mixing different ratios of methylated and non-methylated bisulfite treated DNA. HRM methylation analysis was performed using Light Cycler®480 Gene Scanning software, Roche Diagnostics GmbH (Mannheim, Germany). Each PCR amplification and HRM profile analysis was performed in triplicate.

    Article Snippet: For PCR amplification, 1 μl of the bisulfite treated DNA from patients, HCT116, DLD-1 cells, or standards, and primers (Additional file , Additional file ) was added to 19 μl of 5 X Hot FIREPol EvaGreen HRM Mix, Solis BioDyne Co. (Tartu, Estonia).

    Techniques: DNA Methylation Assay, Methylation Sequencing, DNA Extraction, Amplification, Modification, Sequencing, Polymerase Chain Reaction, Purification, Clone Assay, Plasmid Preparation, Isolation, Software, Methylation, Real-time Polymerase Chain Reaction

    Ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level in three ranges of PHD3 methylation status: 0–1%; 1–10% and 10–100%. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. The methylation for each patient was calculated as an average percentage of methylation in amplified fragments located in the CpG island of PHD3. The samples were divided into three groups for statistical analysis: 0–1% methylation, 1–10% methylation and 10–100% methylation (Table 2 ) [ 28 - 30 ]. To evaluate the statistically significant difference in the ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level between the three DNA methylation ranges (0–1% methylation, 1–10% methylation and 10–100% methylation), the non-parametric Kruskal-Wallis test was employed.

    Journal: BMC Cancer

    Article Title: Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer

    doi: 10.1186/1471-2407-13-526

    Figure Lengend Snippet: Ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level in three ranges of PHD3 methylation status: 0–1%; 1–10% and 10–100%. Methylation percentage of three DNA fragments within the PHD3 CpG island (Additional file 1 , Additional file 2 ) was determined by Real Time PCR amplification of bisulfite treated standard and patient DNA, followed by comparison of their HRM profiles. The methylation for each patient was calculated as an average percentage of methylation in amplified fragments located in the CpG island of PHD3. The samples were divided into three groups for statistical analysis: 0–1% methylation, 1–10% methylation and 10–100% methylation (Table 2 ) [ 28 - 30 ]. To evaluate the statistically significant difference in the ratio of cancerous PHD3 mRNA level to histopathologically unchanged tissue PHD3 mRNA level between the three DNA methylation ranges (0–1% methylation, 1–10% methylation and 10–100% methylation), the non-parametric Kruskal-Wallis test was employed.

    Article Snippet: For PCR amplification, 1 μl of the bisulfite treated DNA from patients, HCT116, DLD-1 cells, or standards, and primers (Additional file , Additional file ) was added to 19 μl of 5 X Hot FIREPol EvaGreen HRM Mix, Solis BioDyne Co. (Tartu, Estonia).

    Techniques: Methylation, Real-time Polymerase Chain Reaction, Amplification, DNA Methylation Assay

    Effect of 5-dAzaC on PHD3 gene DNA methylation in HCT116 and DLD-1 CRC cells. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions either in the absence or in the presence of 5-dAzaC at a concentration of 5.00 μM for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island: A (chr14: 34 419 922–34 420 080), B (chr14: 34 419 795–34 419 935) and C (chr14: 34 419 400–34 419 538) (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles.

    Journal: BMC Cancer

    Article Title: Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer

    doi: 10.1186/1471-2407-13-526

    Figure Lengend Snippet: Effect of 5-dAzaC on PHD3 gene DNA methylation in HCT116 and DLD-1 CRC cells. HCT116 and DLD-1 cells were cultured under normoxic or hypoxic (1% O 2 ) conditions either in the absence or in the presence of 5-dAzaC at a concentration of 5.00 μM for 48 hrs. Cells were then used for DNA isolation followed by bisulfite modification. Methylation percentage of three DNA fragments within the PHD3 CpG island: A (chr14: 34 419 922–34 420 080), B (chr14: 34 419 795–34 419 935) and C (chr14: 34 419 400–34 419 538) (Additional file 1 , Additional file 2 ) in HCT116 and DLD-1 cells under hypoxic and normoxic conditions was determined by Real Time PCR amplification of bisulfite treated standard and cell line DNA, followed by comparison of their HRM profiles.

    Article Snippet: For PCR amplification, 1 μl of the bisulfite treated DNA from patients, HCT116, DLD-1 cells, or standards, and primers (Additional file , Additional file ) was added to 19 μl of 5 X Hot FIREPol EvaGreen HRM Mix, Solis BioDyne Co. (Tartu, Estonia).

    Techniques: DNA Methylation Assay, Cell Culture, Concentration Assay, DNA Extraction, Modification, Methylation, Real-time Polymerase Chain Reaction, Amplification