MethylAffinity™ Methylated DNA enrichment kit provides a GCM™ bead-based method for quick enrichment of methylated DNA fragments from whole genome. The enriched sample improves the performance of downstream analysis, such as RT-PCR, microarray and sequencing for methylation status and location studies.
The genetically engineered GCM recombinant protein is designed based on the functional domain of methyl binding domain (MBD) proteins. It binds specifically to double-stranded DNA containing methylated CpG dinucleotides and has much higher affinity and specificity than any wild-type MBDs.
Protocol overview
Advantages
Fewer steps and less time
Whole process takes < 2 hours
Immobilized GCM magenta beads are provided. No need to perform beads-protein coupling reaction.
GCM captures double-stranded methylated DNA, which simplifies genomic sample preparation and downstream analysis.
Rapid elution with salt.
High affinity
Higher affinity than other commercially available MBD proteins.
Be able to isolate DNA fragments which contain only a few copies of methylated CpG dinucleotides.
Be able to enrich methylated DNA from nanograms of genomic DNA sample.
Gradient elution option
For precise study of methylation status, salt gradient elution can be used to fractionate DNA fragments based on methylated-CpG density.
Convenience
Vivid magenta color makes the beads easy to spot, thus greatly reduces the risk of incidental beads loss.
Restriction enzymes are provided in the kit for genomic DNA fragmentation.
Figure 1. PCR results of enrichment controls
100 ng of mouse genomic DNA (sonicated) were added to 4 µl of GCM-beads. After 1 hour binding reaction at RT, the beads were washed several times. Then the methylated DNA were eluted from the beads. The wash and elution supernatants (5% of each) were analyzed by PCR using primer sets specific for mouse β-actin promoter (unmethylated), GAPDH promoter (unmethylated), and IAP repeats (methylated). The PCR products were checked by agarose gel electrophoresis.
Figure 2. Easy to spot GCM beads Comparison of GCM magenta beads (right) with conventional Sepharose beads (left). The magenta color of the beads helps reduce accidental beads loss during the handling.
