COBRA-seq: sensitive and quantitative methylome profiling

Hilal Varinli*, Aaron L. Statham, Susan J. Clark, Peter L. Molloy, Jason P. Ross

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

4 Citations (Scopus)
18 Downloads (Pure)

Abstract

Combined Bisulfite Restriction Analysis (COBRA) quantifies DNA methylation at a specific locus. It does so via digestion of PCR amplicons produced from bisulfite-treated DNA, using a restriction enzyme that contains a cytosine within its recognition sequence, such as TaqI. Here, we introduce COBRA-seq, a genome wide reduced methylome method that requires minimal DNA input (0.1–1.0 μg) and can either use PCR or linear amplification to amplify the sequencing library. Variants of COBRA-seq can be used to explore CpG-depleted as well as CpG-rich regions in vertebrate DNA. The choice of enzyme influences enrichment for specific genomic features, such as CpG-rich promoters and CpG islands, or enrichment for less CpG dense regions such as enhancers. COBRA-seq coupled with linear amplification has the additional advantage of reduced PCR bias by producing full length fragments at high abundance. Unlike other reduced representative methylome methods, COBRA-seq has great flexibility in the choice of enzyme and can be multiplexed and tuned, to reduce sequencing costs and to interrogate different numbers of sites. Moreover, COBRA-seq is applicable to non-model organisms without the reference genome and compatible with the investigation of non-CpG methylation by using restriction enzymes containing CpA, CpT, and CpC in their recognition site.

Original languageEnglish
Pages (from-to)1140-1163
Number of pages24
JournalGenes
Volume6
Issue number4
DOIs
Publication statusPublished - 23 Oct 2015

Bibliographical note

Copyright the Author(s) 2015. Version archived for private and non-commercial use with the permission of the author/s and according to publisher conditions. For further rights please contact the publisher.

Keywords

  • CHH
  • COBRA
  • DNA methylation
  • Enhancer
  • Next generation sequencing
  • Non CpG
  • Non-model organism
  • Reduced representation
  • Restriction enzymes

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