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5-Methylcytosine is a prominent epigenetic DNA modification which plays an important role in regulation of gene expression.
The Thermo Scientific EpiJET DNA Methylation Analysis Kit (MspI/HpaII) uses the MspI and HpaII restriction enzymes to analyze DNA methylation status at a specific locus. Epi MspI and Epi HpaII are isoschizomers with differing sensitivities to CpG methylation. When the internal CpG in the 5’-CCGG-3’ tetranucleotide sequence is methylated, cleavage with Epi HpaII is blocked, but cleavage with Epi MspI is not affected.
The Epi MspI and Epi HpaII enzymes are specially formulated for epigenetics studies to complete genomic DNA digestion in 1 hour.
DNA sample quality is important for efficient digestion by restriction endonucleases or downstream applications. We recommend use of spin column based kits, such as the Thermo Scientific GeneJET Genomic DNA Purification Kit (#K0721, #K0722).
Parallel digestion of methylated and unmethylated DNA with the restriction enzyme pair (MspI/HpaII) having different sensitivity to methylation.
Methylated and unmethylated linearized plasmid DNA (pUC19/SmaI) were mixed with human blood genomic DNA and digested in parallel with methylation-sensitive Epi HpaII restriction enzyme and methylation insensitive isoschizomer Epi MspI for 1 hour at 37°C. Methylated plasmid DNA and naturally methylated gDNA were protected from cleavage with Epi HpaII, but were cleaved with insensitive Epi MspI. Unmethylated plasmid DNA was cleaved with both enzymes.
1, 4 – undigested DNA (control)
2, 5 – after digestion with Epi HpaII
3, 6 – after digestion with Epi MspI