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. 2019 Oct 16;9:14864. doi: 10.1038/s41598-019-51423-2

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Schematic representation of MCSeEd. Two samples (e.g., control, stressed samples) (a) are subjected to double digestion (b) with a methylation-insensitive (MseI) and a methylation-sensitive enzyme (AciI, PstI, EcoT22, DpnII; see Supplementary Table S1) and ligated (c) with specific adapters: a Y adapter for the MseI site and a barcoded adapter specific for the methylation-sensitive enzyme (c). After size selection, the fragments are pooled and amplified with adapter-specific primers (MseI primers carry library-specific indices) and sequenced using the 1 × 150 Illumina chemistry (d). Demultiplexed reads (e) are analyzed with the design-appropriate MCSeEd pipeline.