Australian Research Council Centres of Excellence in Plant Energy Biology and Plants for Space, School of Molecular Sciences, The University of Western Australia, Crawley, WA, Australia.
Harry Perkins Institute of Medical Research, Nedlands, WA, Australia.
Methods Mol Biol. 2024;2842:391-403. doi: 10.1007/978-1-0716-4051-7_20.
DNA methylation is a covalent modification of DNA that plays important roles in processes such as the regulation of gene expression, transcription factor binding, and suppression of transposable elements. The use of whole-genome bisulfite sequencing (WGBS) enables the genome-wide identification and quantification of DNA methylation patterns at single-base resolution and is the gold standard for the analysis of DNA methylation. However, the computational analysis of WGBS data can be particularly challenging, as many computationally intensive steps are required. Here, we outline step-by-step an approach for the analysis and interpretation of WGBS data. First, sequencing reads must be trimmed, quality-checked, and aligned to the genome. Second, DNA methylation levels are estimated at each cytosine position using the aligned sequence reads of the bisulfite-treated DNA. Third, regions of differential cytosine methylation between samples can be identified. Finally, these data need to be visualized and interpreted in the context of the biological question at hand.
DNA 甲基化是 DNA 的一种共价修饰，在基因表达调控、转录因子结合和转座元件抑制等过程中发挥着重要作用。全基因组亚硫酸氢盐测序（WGBS）的应用使得在单碱基分辨率下对 DNA 甲基化模式进行全基因组鉴定和定量成为可能，是 DNA 甲基化分析的金标准。然而，WGBS 数据的计算分析可能特别具有挑战性，因为需要许多计算密集型步骤。在这里，我们逐步概述了 WGBS 数据分析和解释的方法。首先，必须对测序reads 进行修剪、质量检查，并与基因组进行比对。其次，使用经亚硫酸氢盐处理的 DNA 的比对序列reads 来估计每个胞嘧啶位置的 DNA 甲基化水平。第三，可以识别样本间差异的胞嘧啶甲基化区域。最后，需要根据当前生物学问题的背景对这些数据进行可视化和解释。
