EZ DNA Methylation kit

DNA methylation profiling Gene specific profiling - whole blood (human) FKBP5

Experiment
DNA methylation profiling Gene specific profiling - whole blood (human) FKBP5
Product
EZ DNA Methylation kit from Zymo Research
Manufacturer
Zymo Research

Protocol tips

Upstream tips
For best results, the CT Conversion Reagent should be prepared freshly and used immediately following preparation.

Publication protocol

We optimized the amplifications of 28 regions covering 302 CpGs within GR and/or CTCF binding sites as well as the transcription start site of the FKBP5 locus (see Additional file 9: Table S4 for primers and mapping of the amplicons). In order to reduce cost and maximize the number of samples per sequencing run, triplicate bisulfite treatments were performed for each sample and then pooled to run one PCR amplification per amplicon [25]. Overall, 200 ng to 500 ng of DNA was used per sample and bisulfite treated using the EZ DNA Methylation Kit (Zymo Research, Irvine, CA). Twenty nanograms of bisulfite-converted DNA was then used for each PCR amplification employing Takara EpiTaq HS Polymerase (Clontech, Saint-Germain-en-Laye, France) and 49 amplification cycles. PCR amplicons were then quantified with the Agilent 2200 TapeStation (Agilent Technologies, Waldbronn, Germany) and pooled in equimolar quantities for each sample. AMPure XP beads (Beckman Coulter, Krefeld, Germany) were used for a double size selection (200–500 bp) to remove primer dimers and high molecular DNA fragments.

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Discussion

Discussion

1 year ago

Author: Jody Hancock Canada

How can I preserve leukocytes for DNA methylation profiling?

I would like to preserve leukocytes for future epigenetic analysis. How can I preserve them effectively in order to perform DNA methylation profiling at a later time?

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Papers

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Manufacturer protocol

Download the product protocol from Zymo Research for EZ DNA Methylation kit below.

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Videos

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