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    Thermo Fisher sybr master mix
    Differential enrichment of H3K9/14ac and H3K4me3 histone modifications at the VPAC1 promoter A. Flow cytometric analysis of mouse CD4 + T cells and CD45R + B cells. Left Panel : Non-adherent splenocytes were stained with anti-CD4-FITC pre- (red line) and post- (blue line) CD4 positive magnetic bead separation ( Materials and Methods ) with typical purities ≥93% (n=3). Right Panel : CD4 depleted non-adherent splenocytes were subsequently stained pre- (red line) and post- (blue line) CD45R-FITC positive magnetic bead separation ( Materials and Methods ) with typical purities ≥97% (n=3). B. Bar graph for <t>Taqman</t> qPCR measurements for VPAC1 from indicated cell type. A hash mark indicates a shift in the y-axis. Data is represented as means +/- SEM for relative VPAC1 levels normalized to HPRT and calculated by 2 (-ΔCt) formula from three independent experiments. The asterisk (*) represents a p≤ 0.05 as compared to CD4 + T cells. C-D. ChIP analysis was performed using purified CD4 + T and CD45R + B cells followed by quantitative <t>SYBR</t> green PCR using VPAC1 primer set 2 or genomic negative control. C. Amplification reactions from SYBR green qPCR using primer set 2 were separated on a 3% agarose gel, stained with ethidium bromide and visualized by a CCD camera (Syngene). This experiment was repeated twice with similar results. D. Bar graph showing fold-enrichment over IgG control of H3K9/14ac and H3K4me3 from the indicated cell types. Cycle thresholds (Ct) were subtracted from input Ct values to obtain relative ΔCt values. Fold increases were calculated by 2 -(ΔCt) formula with non specific IgG levels normalized to input arbitrarily set to 1 (not shown). Data is presented as means +/- SEM from three biologically independent experiments (*, p≤0.05).
    Sybr Master Mix, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 8303 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Differential enrichment of H3K9/14ac and H3K4me3 histone modifications at the VPAC1 promoter A. Flow cytometric analysis of mouse CD4 + T cells and CD45R + B cells. Left Panel : Non-adherent splenocytes were stained with anti-CD4-FITC pre- (red line) and post- (blue line) CD4 positive magnetic bead separation ( Materials and Methods ) with typical purities ≥93% (n=3). Right Panel : CD4 depleted non-adherent splenocytes were subsequently stained pre- (red line) and post- (blue line) CD45R-FITC positive magnetic bead separation ( Materials and Methods ) with typical purities ≥97% (n=3). B. Bar graph for Taqman qPCR measurements for VPAC1 from indicated cell type. A hash mark indicates a shift in the y-axis. Data is represented as means +/- SEM for relative VPAC1 levels normalized to HPRT and calculated by 2 (-ΔCt) formula from three independent experiments. The asterisk (*) represents a p≤ 0.05 as compared to CD4 + T cells. C-D. ChIP analysis was performed using purified CD4 + T and CD45R + B cells followed by quantitative SYBR green PCR using VPAC1 primer set 2 or genomic negative control. C. Amplification reactions from SYBR green qPCR using primer set 2 were separated on a 3% agarose gel, stained with ethidium bromide and visualized by a CCD camera (Syngene). This experiment was repeated twice with similar results. D. Bar graph showing fold-enrichment over IgG control of H3K9/14ac and H3K4me3 from the indicated cell types. Cycle thresholds (Ct) were subtracted from input Ct values to obtain relative ΔCt values. Fold increases were calculated by 2 -(ΔCt) formula with non specific IgG levels normalized to input arbitrarily set to 1 (not shown). Data is presented as means +/- SEM from three biologically independent experiments (*, p≤0.05).

    Journal: Regulatory peptides

    Article Title: A Transcriptionally Permissive Epigenetic Landscape at the Vasoactive Intestinal Peptide Receptor-1 Promoter Suggests an Euchromatin Nuclear Position in Murine CD4 T Cells

    doi: 10.1016/j.regpep.2009.08.010

    Figure Lengend Snippet: Differential enrichment of H3K9/14ac and H3K4me3 histone modifications at the VPAC1 promoter A. Flow cytometric analysis of mouse CD4 + T cells and CD45R + B cells. Left Panel : Non-adherent splenocytes were stained with anti-CD4-FITC pre- (red line) and post- (blue line) CD4 positive magnetic bead separation ( Materials and Methods ) with typical purities ≥93% (n=3). Right Panel : CD4 depleted non-adherent splenocytes were subsequently stained pre- (red line) and post- (blue line) CD45R-FITC positive magnetic bead separation ( Materials and Methods ) with typical purities ≥97% (n=3). B. Bar graph for Taqman qPCR measurements for VPAC1 from indicated cell type. A hash mark indicates a shift in the y-axis. Data is represented as means +/- SEM for relative VPAC1 levels normalized to HPRT and calculated by 2 (-ΔCt) formula from three independent experiments. The asterisk (*) represents a p≤ 0.05 as compared to CD4 + T cells. C-D. ChIP analysis was performed using purified CD4 + T and CD45R + B cells followed by quantitative SYBR green PCR using VPAC1 primer set 2 or genomic negative control. C. Amplification reactions from SYBR green qPCR using primer set 2 were separated on a 3% agarose gel, stained with ethidium bromide and visualized by a CCD camera (Syngene). This experiment was repeated twice with similar results. D. Bar graph showing fold-enrichment over IgG control of H3K9/14ac and H3K4me3 from the indicated cell types. Cycle thresholds (Ct) were subtracted from input Ct values to obtain relative ΔCt values. Fold increases were calculated by 2 -(ΔCt) formula with non specific IgG levels normalized to input arbitrarily set to 1 (not shown). Data is presented as means +/- SEM from three biologically independent experiments (*, p≤0.05).

    Article Snippet: SYBR green master mix and Taqman master mix were obtained from Applied Biosystems.

    Techniques: Flow Cytometry, Staining, Real-time Polymerase Chain Reaction, Chromatin Immunoprecipitation, Purification, SYBR Green Assay, Polymerase Chain Reaction, Negative Control, Amplification, Agarose Gel Electrophoresis

    Gene expression analysis of fatty acid-metabolizing enzymes in control and H-RasV12 expressing fibroblasts by qRT-PCR. ( A ) Gene expression analysis of desaturases ( SCD and FADS ), elongases ( ELOVL ) and acyl-coenzyme A synthetases ( ACSL ). ( B ) Gene expression analysis of phospholipases A2 ( PLA2 ). Ten ng of each cDNA were used as template. Reactions were performed in triplicate, using SYBR green technology StepOne RT-PCR machine to detect amplification. The GAPDH gene was used as endogenous control. The fold expression in H-RasV12 fibroblasts with respect to control is displayed, expressed as Relative Quantity (RQ). The analysis was repeated three times in triplicate and the mean ± S.D. is reported (* p

    Journal: International Journal of Molecular Sciences

    Article Title: Oncogenic H-Ras Expression Induces Fatty Acid Profile Changes in Human Fibroblasts and Extracellular Vesicles

    doi: 10.3390/ijms19113515

    Figure Lengend Snippet: Gene expression analysis of fatty acid-metabolizing enzymes in control and H-RasV12 expressing fibroblasts by qRT-PCR. ( A ) Gene expression analysis of desaturases ( SCD and FADS ), elongases ( ELOVL ) and acyl-coenzyme A synthetases ( ACSL ). ( B ) Gene expression analysis of phospholipases A2 ( PLA2 ). Ten ng of each cDNA were used as template. Reactions were performed in triplicate, using SYBR green technology StepOne RT-PCR machine to detect amplification. The GAPDH gene was used as endogenous control. The fold expression in H-RasV12 fibroblasts with respect to control is displayed, expressed as Relative Quantity (RQ). The analysis was repeated three times in triplicate and the mean ± S.D. is reported (* p

    Article Snippet: 4.8. qRT-PCR RNA was extracted and retro-transcribed as previously described [ ]. cDNA was used to determine the expression of genes listed in . cDNA was used to determine transcript levels by qRT-PCR in a StepOne RT-PCR machine (Applied Biosystems, Foster City, CA, USA) using SYBR® Select Master Mix (Life Technologies, Carlsbad, CA, USA).

    Techniques: Expressing, Quantitative RT-PCR, SYBR Green Assay, Step One RT-PCR, Amplification