Genomic DNA and methylated DNA standards (Cat. #55008; Active Motif) were heat- denatured in 0.4 M NaOH (Sigma) & 10 mM EDTA (Sigma) at 100 °C for 10 min and chilled on ice for 5 min. Samples were two-fold serially diluted and applied to a positively charged nylon membrane (Biodyne B; Pall Life Sciences) using the Bio-dot microfiltration apparatus (Biorad). Membranes were UV-crosslinked at 12,0000 μJ/cm2 (CL1000; UVP), and blocked in TBS, 0.1% Tween-20 (TBST), 5% milk for 1 hr. Primary antibodies against 5-mC (1:1000; cat. 39649; Active Motif) were incubated with the membrane in TBST overnight. Dot blots were washed 3X in TBST, incubated in HRP-conjugated IgG secondary antibodies (GE healthcare) for 1hr, and detected by standard ECL methods (Supersignal West Pico; Thermo Fisher Scientific).
Biodyne b
Manufactured by Pall Corporation
The Biodyne B is a type of lab equipment designed for use in various scientific and research applications. It is a membrane-based product that serves as a support for a range of biomolecules, including proteins, nucleic acids, and other substances. The Biodyne B provides a versatile platform for various laboratory procedures, but a detailed description of its core function without interpretation or extrapolation is not available.
Automatically generated - may contain errors
Lab products found in correlation
Bio dot microfiltration apparatus, by Bio-Rad (2 mentions)
Hrp conjugated igg secondary antibodies, by GE Healthcare (2 mentions)
Supersignal west pico, by Thermo Fisher Scientific (2 mentions)
Ethylene diamine tetraacetic acid (edta), by Merck Group (2 mentions)
Sodium hydroxide (naoh), by Merck Group (2 mentions)
3 protocols using biodyne b
1
Quantitative DNA Methylation Analysis
2
cDNA Gel Blotting and Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
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cDNA fragments were separated on 2% NuSieve 3:1 agarose gel. The gel was treated with denaturation buffer consisting of 1.5 M NaCl and 0.5 M NaOH for 30 min. After neutralizing by treatment with neutralizing buffer (pH 7.5) consisting of 0.5 M Tris-HCl and 3 M NaCl for 30 min, cDNA fragments were transferred to Biodyne B (Pall Life Sciences) using blotting buffer consisting of 3 M NaCl and 8 mM NaOH. cDNA fragments were detected using [33P]-labeled oliginucleotide KO197.
3
Quantitative DNA Methylation Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Genomic DNA and methylated DNA standards (Cat. #55008; Active Motif) were heat- denatured in 0.4 M NaOH (Sigma) & 10 mM EDTA (Sigma) at 100 °C for 10 min and chilled on ice for 5 min. Samples were two-fold serially diluted and applied to a positively charged nylon membrane (Biodyne B; Pall Life Sciences) using the Bio-dot microfiltration apparatus (Biorad). Membranes were UV-crosslinked at 12,0000 μJ/cm2 (CL1000; UVP), and blocked in TBS, 0.1% Tween-20 (TBST), 5% milk for 1 hr. Primary antibodies against 5-mC (1:1000; cat. 39649; Active Motif) were incubated with the membrane in TBST overnight. Dot blots were washed 3X in TBST, incubated in HRP-conjugated IgG secondary antibodies (GE healthcare) for 1hr, and detected by standard ECL methods (Supersignal West Pico; Thermo Fisher Scientific).
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