Oral Biology pp 279-298 | Cite as

Generating Multiple Base-Resolution DNA Methylomes Using Reduced Representation Bisulfite Sequencing

  • Aniruddha ChatterjeeEmail author
  • Euan J. Rodger
  • Peter A. Stockwell
  • Gwenn Le Mée
  • Ian M. MorisonEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1537)


Reduced representation bisulfite sequencing (RRBS) is an effective technique for profiling genome-wide DNA methylation patterns in eukaryotes. RRBS couples size selection, bisulfite conversion, and second-generation sequencing to enrich for CpG-dense regions of the genome. The progressive improvement of second-generation sequencing technologies and reduction in cost provided an opportunity to examine the DNA methylation patterns of multiple genomes. Here, we describe a protocol for sequencing multiple RRBS libraries in a single sequencing reaction to generate base-resolution methylomes. Furthermore, we provide a brief guideline for base-calling and data analysis of multiplexed RRBS libraries. These strategies will be useful to perform large-scale, genome-wide DNA methylation analysis.

Key words

Epigenetics DNA methylation Reduced representation bisulfite sequencing Multiplexed Second-generation sequencing CpG island DMAP 



We gratefully acknowledge the help and support of Dr. Robert Day, Dr. Rebecca Laurie, and Les McNoe of the Otago Genomics and Bioinformatics Facility (OGBF), Dunedin, New Zealand, during the development of this method. This work was supported by Gravida, National Centre for Growth and Development (formerly NRCGD) and the Health Research Council (HRC), New Zealand. A.C. would like to gratefully acknowledge the New Zealand Institute for Cancer Research Trust (NZICRT) for their support.


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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Department of Pathology, Dunedin School of MedicineUniversity of OtagoDunedinNew Zealand
  2. 2.Gravida: National Centre for Growth and DevelopmentUniversity of AucklandGrafton, AucklandNew Zealand
  3. 3.Department of BiochemistryUniversity of OtagoDunedinNew Zealand

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