Methods Used to Study Epigenetics
The most common technique for assessing DNA methylation involves the use of sodium bisulfite to convert unmethylated and methylated cytosine residues to uracil and cytosine, respectively. Methylated cytosines can then be identified through various downstream nucleic acid analysis methods, including PCR, qPCR, HRM, and sequencing. This commonly used technique can both identify individual methylation sites and quantify the level of methylation for a particular genomic region.
Other techniques for methylation analysis include the use of restriction enzymes, which are either resistant or sensitive to DNA methylation, and methylated DNA immunoprecipitation (MeDIP), which utilizes antibodies to isolate methylated DNA fragments for analysis.
The interaction between proteins (for example, histones) and DNA is most often studied using a technique known as chromatin immunoprecipitation (ChIP). ChIP is a preparatory method that uses highly specific antibodies against DNA-binding proteins to isolate DNA fragments that bind transcription factors or other DNA-binding proteins.
The isolated fragment can be characterized by using various nucleic acid analysis techniques, including PCR, qPCR, sequencing, and microarray hybridization. This can help determine whether specific proteins are associated with specific genomic regions. ChIP is also useful for identifying regions of the genome that are associated with specific histone modifications (for example, activating or repressive).
Researchers are interested in understanding the role of epigenetic changes, including DNA methylation and histone modifications, in disease (cancer) development and cell differentiation and reprogramming. The methods described thus far provide detailed molecular information about epigenetic markers that can be correlated to changes in gene-expression levels. However, they do not provide direct information regarding the chromatin state associated with these epigenetic marks.
Furthermore, several of these methods are time-consuming and may require up to five days to complete. New tools that deliver additional information about the functional state of chromatin in a shorter time are needed to accelerate the pace of epigenetics research.