Don’t want to chop up your DNA with enzymes, or blast it with bisulfite? Why not try an enrichment-based approach. Enrichment methods enrich a sample for methylated DNA and compare the sequences obtained between experimental conditions.
Methylated DNA immunoprecipitation (MeDIP) has evolved from ChIP. MeDIP uses an antibody against methylcytosine to enrich for methylated regions (Jacinto et al., 2008). Analysis is then very similar to ChIP i.e. the samples can be run on microarray or next-gen sequencing.
Methyl-CpG-binding domain (MBD) capture works differently, it uses beads coated in MBD polypeptide that bind methylated DNA from a genomic sample. After enrichment in this way, the downstream analysis is very similar to MeDIP.
Both strategies have distinct advantages over bisulfite. For one, they will not detect 5hmC as bisulfite does. Further, they do not damage the DNA like bisulfite treatment. Between MeDIP and MBD capture, MeDIP is more sensitive to methylation differences in regions of low CpG density, while MBD is more sensitive in regions with higher CpG density such as CpG islands (Nair et al., 2011).
In both techniques, a necessary whole-genome amplification step prior to array/sequencing can introduce a bias against CpG sites (Robinson et al., 2010). Other methods such as Agilent SureSelect uses microarray- or solution-capture or DNA regions that have important methylations (CpG islands, imprinted regions etc.). They rely on sequence information to pull out these regions, not methylation.
DNA Methylation Enrichment Additional Reading
This paper presents the MeDIP-chip experimental design and focuses on quality control and data analysis hurdles.
This paper describes the MBD capture (aka MethylCap-Seq) protocol including data analysis an interpretation. The authors pay special attention to comparing the outcome of varying different experimental conditions.
- Jacinto, F.V., Ballestar, E., and Esteller, M. (2008). Methyl-DNA immunoprecipitation (MeDIP): hunting down the DNA methylome. BioTechniques 44, 35, 37, 39 passim.
- Nair, S.S., Coolen, M.W., Stirzaker, C., Song, J.Z., Statham, A.L., Strbenac, D., Robinson, M.D., and Clark, S.J. (2011). Comparison of methyl-DNA immunoprecipitation (MeDIP) and methyl-CpG binding domain (MBD) protein capture for genome-wide DNA methylation analysis reveal CpG sequence coverage bias. Epigenetics 6, 34-44.
- Robinson, M.D., Stirzaker, C., Statham, A.L., Coolen, M.W., Song, J.Z., Nair, S.S., Strbenac, D., Speed, T.P., and Clark, S.J. (2010). Evaluation of affinity-based genome-wide DNA methylation data: effects of CpG density, amplification bias, and copy number variation. Genome Res. 20, 1719-1729.