EpiTYPER DNA Methylation analysis
There are four steps in the EpiTYPER DNA Methylation analysis, including bisulfite conversion, PCR, MassCLEAVE (T7-IVT and Base-specific RNA cleavage) and Mass detection.
- The unmethylated cytosines in the DNA sequence are converted to uridines after the bisulfite treatment.
- Then T7 RNA polymerase will bind to the T7 RNA polymerase promoter sequence on the reverse PCR primer and perform in vitro transcription and transfer the CTP into dCTP.
- In the meantime, the single strand RNA product will be cleaved by RNase A during the MassCLEAVE process.
- The whole sequence will be cleaved to tens of fragments generally. The fragment that contained CpG sites will have two kinds of molecular weight (GC or AG) because of the methylation status. There are 16 Daltons of discrepancy between the methylated and unmethylated sequence. We can report the methylation ratio data base on the signal area.