EpiQuik Dnmt3A Assay Kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	gDNA was extracted using the NucleoSpin® Tissue kit (740952, Macherey Nagel). Global DNA methylation was quantified using the MethylFlash methylated DNA quantification kit (P-1034, Epigentek, France). Methyl DNA collection were performed using the “Methyl-Collector Ultra kit” (55005, Active Motif). Histone 3 acetylation was quantified using the EpiQuik Tissue Acetyl-Histone H3 ChIP Kit (P-2012, Epigentek). ChIP was performed using the ChIP-IT High Sensitivity kit (53040, Active Motif) with ChIP grade antibodies (Table ). All these kits were used according to the manufacturer’s instructions.

Protocol tips

gDNA was extracted using the NucleoSpin® Tissue kit (740952, Macherey Nagel). Global DNA methylation was quantified using the MethylFlash methylated DNA quantification kit (P-1034, Epigentek, France). Methyl DNA collection were performed using the “Methyl-Collector Ultra kit” (55005, Active Motif). Histone 3 acetylation was quantified using the EpiQuik Tissue Acetyl-Histone H3 ChIP Kit (P-2012, Epigentek). ChIP was performed using the ChIP-IT High Sensitivity kit (53040, Active Motif) with ChIP grade antibodies (Table ). All these kits were used according to the manufacturer’s instructions.

Publication protocol

DNMT quantification
Quantification of nuclear DNMT3A and DNMT3B enzyme concentrations were performed as per the manufacturer's guidelines (Epiquik DNMT3A/B Assay Kit, Epigentek, New York). All samples were assayed in duplicate, and various dilutions of the respective DNMT standards were used to plot a standard curve. Microplates were scanned at 450 nm with a reference wavelength of 655 nm. Intra‐assay coefficient of variations for DNMT3A and DNMT3B were calculated as 3.95% and 9.92%, respectively.

The following equation was used to quantify enzyme concentration:

Concentration(ng · mg of protein-1) = (sample OD - blank OD)/(slope × protein amount) × 1000
where OD is optical density/absorbance. Slope is generated from the standard curve. Protein amount (μg) as quantified by Bradford assay.

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Manufacturer protocol

Download the product protocol from Epigentek for EpiQuik Dnmt3A Assay Kit below.

Download manufacturer protocol

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