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DNA methylation was the first epigenetic mark to be discovered, involving the addition of a methyl group to the 5 position of cytosine by DNA methyltransferases, and can be inherited through cell division. DNA hydroxymethylation, caused by oxidation of 5-methylcytosine through the TET family of enzymes, was further discovered to be involved in switching genes on and off. DNA methylation plays an important role in normal human development and is associated with the regulation of gene expression, tumorigenesis, and other genetic and epigenetic diseases.
Bisulfite DNA Modification
Also known as bisulfite conversion, are common approaches for preparing DNA for gene-specific DNA methylation analysis. Treatment of DNA with bisulfite converts cytosine to uracil while leaving 5-methylcytosine intact, allowing for single-nucleotide resolution information about the methylated areas of DNA. Popular methods for analyzing DNA methylation on a gene-specific basis, after bisulfite treatment, include bisulfite sequencing, methylation specific PCR, and methylation-based microarrays.Chemical-based DNA bisulfite conversion
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Methylamp DNA Modification Kit |
Methylamp One-Step DNA Modification Kit |
BisulFlash DNA Modification Kit |
Methylamp Whole Cell Bisulfite Modification Kit |
Methylamp 96 DNA Modification Kit |
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| Purpose |
Chemical-based DNA bisulfite conversion |
Heat-based DNA bisulfite conversion |
Fast heat-based DNA bisulfite conversion |
Directly convert from cells, tissues, saliva, urine, or blood |
High-throughput 96-well format bisulfite conversion |
| Lowest Input Amount |
50 pg |
50 pg |
200 pg |
100 cells, 1 µl blood |
100 pg |
| Total Protocol Time |
< 2 hours |
< 2 hours |
30 minutes |
< 3 hours |
< 2 hours 30 minutes |
| Elution Volume |
8-20 µl |
8-20 µl |
10-20 µl |
8-20 µl |
20-40 µl |
| Conversion Efficiency |
99.9% |
99.5% |
99.9% |
99.9% |
99.5% |
| Desulfonation / Clean-Up |
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| Final Yield |
>75% |
>75% |
>75% |
>85% |
>75% |
| Post-Conversion DNA Fragment Size |
200-2000 bp; Avg. Peak of 800 bp |
200-2000 bp; Avg. Peak of 800 bp |
100-400 bp; Avg. Peak of 250 bp |
200-2000 bp; Avg. Peak of 800 bp |
200-2000 bp; Avg. Peak of 800 bp |
| Vessel Format |
Spin Column |
Spin Column |
Spin Column |
Spin Column |
96-Well Plate |
DNA Methylation Quantification
DNA methylation plays an important role in normal organismal development and in cellular differentiation in higher organisms. Gene expression, as well as the development of nearly all types of cancer, are also tied to DNA methylation. For example, global decrease in 5-methylcytosine content (DNA hypomethylation) is likely caused by methyl-deficiency due to a variety of environmental influences, and has been proposed as a molecular marker in multiple biological processes such as cancer. Global quantification of DNA methylation is crucial for understanding the roles that gene expression and silencing play in the development of cancer and other diseases.
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MethylFlash Methylated DNA Quantification Kit (Colorimetric) |
MethylFlash Urine 5-Methylcytosine (5-mC) Quantification Kit (Colorimetric) |
MethylFlash Hydroxymethylated DNA Quantification Kit (Colorimetric) |
MethylFlash 5-Formylcytosine (5-fC) DNA Quantification Kit (Colorimetric) |
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| Purpose |
Colorimetrically measure global 5mC in DNA |
Colorimetrically measure global 5mC in DNA directly from urine |
Colorimetrically measure global 5hmC in DNA |
Colorimetrically measure global 5fC in DNA |
| Input Range |
50-200 ng |
0.5-5.0 µl |
50-200 ng |
100-300 ng |
| Optimal Input Amount |
100 ng |
2 µl |
100 ng |
300 ng |
| Total Time |
4 hours |
4 hours |
4 hours |
4 hours |
| Detection Threshold |
200 pg of meDNA |
30 pg of meDNA |
40 pg of hMeDNA |
1 pg of fC DNA |
| Specificity |
>1000:1 meDNA:unMeDNA |
>1000:1 meDNA:unMeDNA |
>1000:1 hMeDNA:unMeDNA |
>1000:1 fC DNA:unMeDNA |
| Format |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
Methylated DNA Immunoprecipitation
Methylated DNA immunoprecipitation (meDIP) is a large scale antibody-based technique that is used to enrich and capture methylated DNA fragments for use in gene-specific DNA methylation studies on a genome wide scale. Following meDIP approaches, DNA methylation can be analyzed using a variety of downstream applications including meDIP-PCR, meDIP-ChIP, and meDIP-sequencing.
DNA Methyltransferase & Demethylase Assays
DNA methyltransferases, or DNMTs, catalyze DNA methylation by adding methyl groups to the 5-carbon position of the cystosine ring, resulting in 5-methylcytosine. The various types of DNMTs are responsible for the maintenance and establishment of DNA methylation patterns. The ten-eleven translocation, or TET, family of 5-mC hydroxylase enzymes including TET1 are responsible for oxidizing 5-methylcytosine into 5-hydroxymethylcytosine. Measuring DNMT and TET amounts, activity, and inhibition levels will allow us to better understand the relationships between cytosine, 5-methylcytosine, and 5-hydroxymethylcytosine as part of the DNA methylation cycle
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EpiQuik DNMT Activity/Inhibition Assay Ultra Kit (Colorimetric) |
Epigenase 5mC-Hydroxylase TET Activity/Inhibition Assay Kit (Colorimetric) |
Epigenase Thymine DNA Glycosylase (TDG) Activity/Inhibition Assay Kit (Colorimetric) |
EpiQuik DNA Demethylase Activity/Inhibition Assay Ultra Kit |
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| Purpose |
Colorimetrically measure total DNMT activity/inhibition |
Colorimetrically measure total TET activity/inhibition |
Colorimetrically measure total TDG activity/inhibition |
Colorimetrically measure total DNA demethylase activity/inhibition |
| Starting Material |
Nuclear extract or purified enzyme |
Nuclear extract or purified enzyme |
Nuclear extract or purified enzyme |
Nuclear extract |
| Input Range |
0.5-20 µg of nuclear extract; or 20-200 ng of purified enzyme |
2-20 µg of nuclear extract; or 20-1000 ng of purified enzyme |
2-10 µg of nuclear extract; or 20-1000 ng of purified enzyme |
2-20 µg of nuclear extract |
| Total Time |
4 hours |
5 hours |
5 hours |
4 hours |
| Detection Threshold |
0.5 ng of purified DNMT |
20 ng of purified TET |
20 ng of purified TDG |
1 µg of nuclear extract |
| Positive Control Included |
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| Format |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
ELISA-like Microplate-based |
Methylated DNA Amplification
Methylation specific PCR (MSP or MS-PCR) and bisulfite sequencing are some of the most commonly used methods for analyzing gene or sequence-specific DNA methylation data, usually after bisulfite treatment of DNA. After conversion, selective methylation amplification of a target region of methylated sequences with methylation primers is often necessary in methylation analysis protocols.