Lesur Isabelle, Rogier Odile, Sow Mamadou Dia, Boury Christophe, Duplan Alexandre, Garnier Abel, Senhaji-Rachik Abdeljalil, Civan Peter, Daron Josquin, Delaunay Alain, Duvaux Ludovic, Benoit Vanina, Guichoux Erwan, Le Provost Grégoire, Sanou Edmond, Ambroise Christophe, Plomion Christophe, Salse Jérôme, Segura Vincent, Tost Jörg, Maury Stéphane
INRAE, Univ. Bordeaux, BIOGECO, F-33610 Cestas, France.
HelixVenture, F-33700 Mérignac, France.
J Exp Bot. 2024 Sep 27;75(18):5568-5584. doi: 10.1093/jxb/erae266.
In the last 20 years, several techniques have been developed for quantifying DNA methylation, the most studied epigenetic marks in eukaryotes, including the gold standard method, whole-genome bisulfite sequencing (WGBS). WGBS quantifies genome-wide DNA methylation but has several inconveniences rendering it less suitable for population-scale epigenetic studies. The high cost of deep sequencing and the large amounts of data generated prompted us to seek an alternative approach. Restricting studies to parts of the genome would be a satisfactory alternative had there not been a major limitation: the need to select upstream targets corresponding to differentially methylated regions as targets. Given the need to study large numbers of samples, we propose a strategy for investigating DNA methylation variation in natural populations, taking into account the structural complexity of genomes, their size, and their content in unique coding regions versus repeated regions as transposable elements. We first identified regions of highly variable DNA methylation in a subset of genotypes representative of the biological diversity in the population by WGBS. We then analysed the variations of DNA methylation in these targeted regions at the population level by sequencing capture bisulfite (SeqCapBis). The entire strategy was then validated by applying it to another species. Our strategy was developed as a proof of concept on natural populations of two forest species: Populus nigra and Quercus petraea.
在过去20年里，已经开发出了几种用于量化DNA甲基化的技术，DNA甲基化是真核生物中研究最多的表观遗传标记，其中包括金标准方法——全基因组亚硫酸氢盐测序（WGBS）。WGBS可对全基因组的DNA甲基化进行量化，但存在一些不便之处，使其不太适合大规模人群的表观遗传学研究。深度测序的高成本以及产生的大量数据促使我们寻求替代方法。如果不存在一个主要限制因素，将研究局限于基因组的某些部分会是一个令人满意的替代方案：即需要选择与差异甲基化区域相对应的上游靶点作为研究对象。鉴于需要研究大量样本，我们提出了一种在自然种群中研究DNA甲基化变异的策略，该策略考虑了基因组的结构复杂性、大小以及其在独特编码区域与作为转座元件的重复区域中的含量。我们首先通过WGBS在代表种群生物多样性的一部分基因型中确定了DNA甲基化高度可变的区域。然后，我们通过测序捕获亚硫酸氢盐（SeqCapBis）在种群水平上分析了这些靶向区域中DNA甲基化的变异情况。最后，通过将整个策略应用于另一个物种对其进行了验证。我们的策略是作为对两种森林物种（黑杨和岩生栎）自然种群的概念验证而开发的。
