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Bs RAD seq: screening DNA methylation in natural populations of non‐model species
Author(s) -
Trucchi Emiliano,
Mazzarella Anna B.,
Gilfillan Gregor D.,
Lorenzo Maria T.,
Schönswetter Peter,
Paun Ovidiu
Publication year - 2016
Publication title -
molecular ecology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.619
H-Index - 225
eISSN - 1365-294X
pISSN - 0962-1083
DOI - 10.1111/mec.13550
Subject(s) - biology , dna methylation , epigenetics , genetics , bisulfite sequencing , genome , epigenomics , computational biology , methylation , evolutionary biology , dna , gene , gene expression
Epigenetic modifications are expected to occur at a much faster rate than genetic mutations, potentially causing isolated populations to stochastically drift apart, or if they are subjected to different selective regimes, to directionally diverge. A high level of genome‐wide epigenetic divergence between individuals occupying distinct habitats is therefore predicted. Here, we introduce bisulfite‐converted restriction site associated DNA sequencing (bs RAD seq), an approach to quantify the level of DNA methylation differentiation across multiple individuals. This reduced representation method is flexible in the extent of DNA sequence interrogated. We showcase its applicability in three natural systems, each comprising individuals adapted to divergent environments: a diploid plant ( Heliosperma, Caryophyllaceae), a tetraploid plant ( Dactylorhiza, Orchidaceae) and an animal ( Gasterosteusaculeatus, Gasterosteidae). We present a robust bioinformatic pipeline, combining tools for RAD locus assembly, SNP calling, bisulfite‐converted read mapping and DNA methylation calling to analyse bs RAD seq data with or without a reference genome. Importantly, our approach accurately distinguishes between SNP s and methylation polymorphism ( SMP s). Although DNA methylation frequency between different positions of a genome varies widely, we find a surprisingly high consistency in the methylation profile between individuals thriving in divergent ecological conditions, particularly in Heliosperma . This constitutive stability points to significant molecular or developmental constraints acting on DNA methylation variation. Altogether, by combining the flexibility of RAD seq with the accuracy of bisulfite sequencing in quantifying DNA methylation, the bs RAD seq methodology and our bioinformatic pipeline open up the opportunity for genome‐wide epigenetic investigations of evolutionary and ecological relevance in non‐model species, independent of their genomic features.