EZ DNA Methylation Kit
D5001 / D5002
- Streamlined, proven procedure for bisulfite conversion of DNA.
- Desulphonation and recovery of bisulfite-treated DNA with a spin column.
- Recovered DNA is ideal for downstream analyses including PCR, endonuclease digestion, sequencing, microarrays, etc.
|Applications||Purified, converted DNA is of high-quality and well-suited for downstream processes, including library preparation for Next-generation sequencing, PCR amplification, etc. This kit is recommended for use with Illumina Infinium MethylationEPIC BeadChip array.|
|Elution Volume||≥10 µl|
|Equipment||Microcentrifuge and thermocycler with heated lid|
|Input||500 pg - 2 µg of DNA|
|Processing Time||12-16 hours|
|Sample Source||Purified genomic DNA, endonuclease-digested DNA, linearized plasmid DNA, etc. DNA should be high-quality and RNA-free.|
Q1: Is an incubation with desulphonation buffer for longer than 20 minutes recommended?
Q2: Tips for bisulfite primer design?
Q3: Which polymerase is recommended for amplification from bisulfite converted DNA?
Q4: Does bisulfite conversion only occur in a CpG context?
Q5: What is the minimum DNA size that can be recovered?
Q6: How to quantify / visualize converted DNA?
Q7: What leads to poor conversion efficiency/ low yields?
Q8: How long is bisulfite converted DNA stable at -20 °C?
Researchers from Germany used the EZ DNA Methylation Kit to bisulfite convert genomic DNA from ex vivo isolated T cell subsets to study the changes in DNA methylation during differentiation of naïve T cells into Th cell subsets. Further methylome analysis through methylation sensitive high-resolution melting (MS-HRM) and pyrosequencing demonstrated that CD4+ T cells share more demethylated regions with Th17 cells when compared to Th1 cells, and that overall Th17 cells display the highest number of demethylated regions. Through this analysis, the researchers were able to identify a unique Th17-specific epigenetic signature and reinforce the finding that Th cell differentiation is associated with major epigenetic changes.Yang BH, et. al. (2015) Development of a unique epigenetic signature during in vivo Th17 differentiation. Nucleic Acids Res. Feb 18;43(3):1537-48. doi: 10.1093/nar/gkv014. Epub 2015 Jan 15.
In a genome-wide study investigating DNA methylation levels in acute myeloid leukemia cells, researchers used the EZ DNA MethylationTM Kit to bisulfite convert genomic DNA obtained from clinical samples. PCR amplification followed by bisulfite sequencing of the converted DNA helped to show that patients diagnosed with acute myeloid leukemia have distinct methylomes in their cancer cells compared to normal cells.Saied et al. (2012) Genome wide analysis of acute myeloid leukemia reveal leukemia specific methylome and subtype specific hypomethylation of repeats. PLoS One. 7(3):e33213.
In a study to determine how global DNA methylation patterns change with age, researchers used the EZ DNA Methylation Kit to bisulfite convert genomic DNA blood samples taken from newborn babies and 100+ year old patients. The authors subsequently subjected the bisulfite-converted DNA to whole genome bisulfite sequencing, 450 K CpG site microarray analysis, and bisulfite genomic sequencing and pyrosequencing, and they found that genome wide DNA methylation content significantly decreased with age.Heyn H, Li N, Ferreira HJ, et al. (2012) Distinct DNA methylomes of newborns and centenarians. Proc Natl Acad Sci U S A. 109(26):10522-7