determine genome-wide DNA methylation patterns
The reliable identification of differential DNA methylation is important for researchers interested in biomarker identification, as well as for those trying to understand the basis of disease, drug mechanism of action or environmental influences on epigenetics. To help speed research in these areas, Active Motif now offers MeDIP-Seq as an end-to-end, genome-wide epigenetic service to identify differentially methylated regions.
The MeDIP-Seq Service includes:
Customers submit purified DNA, frozen tissues or cell pellets, then we:
- Prepare the sample.
- Perform MeDIP with a 5-mC mAb.
- Amplify the enriched DNA sample.
- Perform Next-Gen sequencing.
- Analyze the data and deliver it to the customer.
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How does MeDIP-Seq work?
In MeDIP-Seq, a highly specific antibody that recognizes 5-methylcytosine is used to immunoprecipitate sonicated genomic DNA, resulting in the enrichment of genomic regions that are methylated. Because 5-methylcytosine antibody binds only to methylated cytosines in the context of single-stranded DNA, the DNA must be denatured prior to immunoprecipitation. As a result of denaturation the enriched DNA can not be processed for Next-Gen sequencing using the typical sequencing library generation protocols, as these require adaptor ligation to double-stranded DNA. This problem is circumvented by ligating the Next-Gen sequencing adaptors to genomic DNA prior to the immunoprecipitation. Following MeDIP, the enriched regions can be directly amplified with Next-Gen sequencing compatible primers. Unique alignment of the sequence tags across the genome reveals the regions of DNA methylation.
Results from Active Motif’s MeDIP-Seq Service correlate well with published MeDIP results.
Several methodologies exist that can detect DNA methylation on a genome-wide scale. Whole genome bisulfite sequencing is the most comprehensive, providing single base resolution across the entire genome. However, it requires sequencing of the entire genome, which is extremely expensive. Reduced Representation Bisulfite Sequencing (RRBS) also yields single base resolution, but it only interrogates 10% of all CpGs and it is more heavily biased toward CpG-rich regions than any other technique. This is not necessarily an advantage as most CpG islands are unmethylated, and because there is growing interest and focus on regions with lower CpG density. MeDIP-Seq coverage is not as biased toward high CpG density and in theory can interrogate all regions across the genome. The expanded genomic coverage of MeDIP-Seq compared to RRBS increases the likelihood of identifying differentially methylated regions in multi-sample studies.
MeDIP-Seq data compared to CpG density and RRBS.