A comprehensive evaluation of alignment software for reduced representation bisulfite sequencing data
A comprehensive evaluation of alignment software for reduced representation bisulfite sequencing data
复制标题
对减少代表性亚硫酸氢盐测序数据的比对软件进行全面评估。
DOI：
10.1093/bioinformatics/bty174
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发表时间：
2018-08-15
期刊：
影响因子：
5.8
通讯作者：
Liu, Pengyuan
中科院分区：
文献类型：
--
作者：
Sun, Xiwei;Han, Yi;Liu, Pengyuan
动机：下一代测序技术的快速发展为单碱基分辨率下研究全基因组DNA甲基化提供了机会。然而，未甲基化胞嘧啶的耗尽为将亚硫酸氢盐转化的测序读数与大的参考比对带来了挑战。用于比对甲基化读段的软件工具尚未得到全面评估，特别是对于广泛使用的涉及富集CpG岛（CGIs）的减少代表性亚硫酸氢盐测序（RRBS）。我们在真实的和模拟的RRBS数据中对7种用于甲基化分析的作图算法进行了广泛的比较。18例肺肿瘤及配对的癌旁组织按RRBS方案测序。我们的经验评估发现，对于低测序深度、中等甲基化水平、CGI岸边或基因体上的CpG位点，软件工具之间的甲基化结果不太一致。这些观察结果通过模拟进一步证实，模拟表明软件工具通常具有检测这些脆弱CpG位点的较低召回率和估计这些CpG位点中甲基化水平的较低精度。在测试的软件工具中，bwa-meth和BS-Seeker 2（bowtie 2）是目前我们在召回率，精度和速度方面首选的RRBS数据对齐器。现有的比对器不能有效地处理中度甲基化的CpG位点和CGI海岸或基因体上的那些CpG位点。对这些易受攻击的CpG位点的甲基化结果的解释应谨慎对待。我们的研究揭示了甲基化数据中固有的几个重要特征，RRBSsim为推进基于序列的甲基化数据分析和方法开发提供了指导。可用性和实施：RRBSsim是RRBS数据基准分析的模拟器，其源代码可在https://github.com/xwBio/RRBSsim或https://github.com/xwBio/Docker-RRBSsim.Contact：yanlu76@zju.edu.cn或pyliu@zju.edu. cn补充信息：补充数据可在Bioinformatics online获得。
Motivation: The rapid development of next-generation sequencing technology provides an opportunity to study genome-wide DNA methylation at single-base resolution. However, depletion of unmethylated cytosines brings challenges for aligning bisulfite-converted sequencing reads to a large reference. Software tools for aligning methylation reads have not yet been comprehensively evaluated, especially for the widely used reduced representation bisulfite sequencing (RRBS) that involves enrichment for CpG islands (CGIs).Results: We specially developed a simulator, RRBSsim, for benchmarking analysis of RRBS data. We performed extensive comparison of seven mapping algorithms for methylation analysis in both real and simulated RRBS data. Eighteen lung tumors and matched adjacent tissues were sequenced by the RRBS protocols. Our empirical evaluation found that methylation results were less consistent between software tools for CpG sites with low sequencing depth, medium methylation level, on CGI shores or gene body. These observations were further confirmed by simulations that indicated software tools generally had lower recall of detecting these vulnerable CpG sites and lower precision of estimating methylation levels in these CpG sites. Among the software tools tested, bwa-meth and BS-Seeker2 (bowtie2) are currently our preferred aligners for RRBS data in terms of recall, precision and speed. Existing aligners cannot efficiently handle moderately methylated CpG sites and those CpG sites on CGI shores or gene body. Interpretation of methylation results from these vulnerable CpG sites should be treated with caution. Our study reveals several important features inherent in methylation data, and RRBSsim provides guidance to advance sequence-based methylation data analysis and methodological development.Availability and implementation: RRBSsim is a simulator for benchmarking analysis of RRBS data and its source code is available at https://github.com/xwBio/RRBSsim or https://github.com/xwBio/Docker-RRBSsim.Contact: yanlu76@zju.edu.cn or pyliu@zju.edu.cnSupplementary information: Supplementary data are available at Bioinformatics online.