Epigenetics

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Facts & Details: 

To study the epigenetic mechanism of DNA methylation we can use the Infinium HumanMethylation450 BeadChip as well as our Reduced Representation Bisulfite Sequencing (RRBS) protocol.

After bisulfite conversion of the samples, the BeadChip quantifies methylation levels at more than 485,000 specific methylation sites per sample at single-nucleotide resolution within the human genome. More details on the array are provided on the vendor’s webpage. The advantage of this BeadChip is that the process is highly automated and, therefore,  hence provides a high-throughput.

Reduced Representation Bisulfite Sequencing (RRBS) uses the restriction enzyme MspI, which cuts specifically at 5´-C^CGG-3´, thereby enriching for sequences containing CpG motives. Following adapter ligation, bisulfite conversion and amplification the RRBS libraries can be sequenced on an Illumina HiSeq machine, providing single-base pair resolution data. In our opinion, RRBS still provides the best cost-benefit trade-off in comparison to whole methylome sequencing, which is still costly, yields 50-fold more reads per sample and is, therefore, computationally more intensive. The following table from Smith et al. (Methods. 2009;48(3):226-32) shows that RRBS covers > 84% of all prominent CpG sites in the mouse and human genome (current read lengths are > 100 bp).

Theoretical coverage of MspI digest fragments based on read length in mouse and human.