Methy-Pipe: An Integrated Bioinformatics Pipeline for Whole Genome Bisulfite Sequencing Data Analysis

DNA methylation, one of the most important epigenetic modifications, plays a crucial role in various biological processes. The level of DNA methylation can be measured using whole-genome bisulfite sequencing at single base resolution. However, until now, there is a paucity of publicly available software for carrying out integrated methylation data analysis. In this study, we implemented Methy-Pipe, which not only fulfills the core data analysis requirements (e.g. sequence alignment, differential methylation analysis, etc.) but also provides useful tools for methylation data annotation and visualization. Specifically, it uses Burrow-Wheeler Transform (BWT) algorithm to directly align bisulfite sequencing reads to a reference genome and implements a novel sliding window based approach with statistical methods for the identification of differentially methylated regions (DMRs). The capability of processing data parallelly allows it to outperform a number of other bisulfite alignment software packages. To demonstrate its utility and performance, we applied it to both real and simulated bisulfite sequencing datasets. The results indicate that Methy-Pipe can accurately estimate methylation densities, identify DMRs and provide a variety of utility programs for downstream methylation data analysis. In summary, Methy-Pipe is a useful pipeline that can process whole genome bisulfite sequencing data in an efficient, accurate, and user-friendly manner. Software and test dataset are available at http://sunlab.lihs.cuhk.edu.hk/methy-pipe/.DNA甲基化是最重要的表观遗传修饰（epigenetic modifications）之一，在多种生物学过程中发挥关键调控作用。采用单碱基分辨率的全基因组亚硫酸氢盐测序（whole-genome bisulfite sequencing）可检测DNA甲基化水平，但截至目前，公开可用的整合式甲基化数据分析软件仍十分匮乏。本研究开发了Methy-Pipe分析流程，其不仅可满足核心数据分析需求（如序列比对、差异甲基化分析等），还提供了实用的甲基化数据注释与可视化工具。具体而言，该工具采用Burrow-Wheeler变换（Burrow-Wheeler Transform, BWT）算法，可直接将亚硫酸氢盐测序读段（bisulfite sequencing reads）比对至参考基因组，并结合统计方法实现了一种基于滑动窗口的新型方法，用于识别差异甲基化区域（differentially methylated regions, DMRs）。其并行数据处理能力使其性能优于多款其他亚硫酸氢盐比对软件。为验证其实用性与性能，本研究将其应用于真实及模拟的亚硫酸氢盐测序数据集。结果表明，Methy-Pipe可准确估算甲基化密度、识别差异甲基化区域，并提供多种实用程序用于下游甲基化数据分析。总而言之，Methy-Pipe是一款高效、准确且用户友好的全基因组亚硫酸氢盐测序数据分析流程。相关软件与测试数据集可通过http://sunlab.lihs.cuhk.edu.hk/methy-pipe/获取。