Important genetic alterations that do not impact DNA sequence

Unlike genetic variants such as SNPs, indels and structural re-arrangements, epigenetic modifications impact gene expression, regulation and/or function without changing the underlying DNA sequence. Epigenetic mechanisms allow cells to respond to their environment by altering phenotypes without a change in genotype. Epigenetic modifications are heritable, often reversible, and play a key role in human development, health and disease. The predominant epigenetic mechanisms are DNA methylation, chromatin modification (e.g. histone acetylation and deacetylation), and non-coding RNAs (particularly micro RNAs).1,2

NGS methods for studying epigenetic modifications

Epigenomics focuses on the analysis of the epigenome, or all of the epigenetic modifications in particular cell or organism. Whilst the molecular basis for epigenetic changes has been studied for several decades,2 high-throughput, next-generation sequencing assays have epigenomic studies on a scale not possible with older molecular technologies.3

  • DNA methylation is now broadly studied using bisulfite sequencing (also known as Methyl-Seq), in which non-methylated cytosine residues are converted to uracils by bisulfite treatment prior to sequencing. A wide array of Methyl-Seq techniques and protocols exist. These include whole-genome bisulfite sequencing (WGBS), targeted bisulfite sequencing, reduced representation bisulfite sequencing (RRBS), and more recently, also single-cell bisulfite sequencing (scBS).3
  • Chromatin modifications are studied using a variety of techniques. Histone modifications are commonly analyzed using ChIP-Seq, in which chromatin immunoprecipitation is followed by NGS. Chromatin accessibility may be studied using nuclease-based assays such as DNAse-Seq and MNase-Seq; FAIRE-Seq (Formaldehyde-Assisted Isolation of Regulatory Elements)4; or transposon-based ATAC-Seq.5

NGS Sample prep for Epigenomics

Methods used in epigenomic studies vary very widely in terms of the way samples are collected, how the nucleic acids of interest are isolated, and how libraries are prepared for NGS. These methods largely have two things in common: very low amounts of the nucleic acid of interest is typically available for library preparation, and one or more PCR amplification steps are required during the library construction process. Chemical processes used for bisulfite conversion and cross-linking are destructive, and render PCR amplification particularly challenging; by either significantly increasing the AT-content of the template, or introducing single- and double-stranded breaks, crosslinked and abasic sites and other molecular damage. For these reasons, highly efficient amplification enzymes and library preparation methods are required to convert precious input DNA to sequencing-ready libraries. Roche Sample Prep Solutions offer engineered enzymes and proven library preparation kits that allow you to process more samples for your epigenomic studies successfully, get more information from every sample, and optimize your sequencing resources.


  1. Tollefsbol T. Translational Epigenetics, Epigenetics in Human Disease. 2nd edition. London:Acadmic Press, 2018. Chapter 1. doi.org/10.1016/B978-0-12-812215-0.00001-7.
  2. https://www.whatisepigenetics.com/fundamentals/. Accessed January 2019.
  3. Masser DR et al. Analysis of DNA modifications in aging research. GeroScience 2018;40:11. doi.org/10.1007/s11357-018-0005-3.
  4. Furey TS. ChIP-seq and beyond: new and improved methodologies to detect and characterize protein-DNA interactions. Nat. Genet. 2012;13:840. doi:10.1038/nrg3306.
  5. Buenrostro JD et al. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat. Methods 2013;10:1213. doi:10.1038/nmeth.2688