Development Of Bioinformatics Tools For DNA Methylation Analysis Based On Next-generation Sequencing

DNA methylation, as an important epigenetic modification, plays an crucial role in X-chromosome inactivation, embryonic development, cell differentiation, disease and cancer formation and other diseases. With the advent of high-throughput sequencing, researches on DNA methylation were facilitated both in the depth and scope, and a large number of genome-wide methylation analysis methods appeared, such as whole-genome bisulfite sequencing (BS-seq), methylated DNA immunoprecipitation sequencing (MeDIP-seq) and reduced representation bisulphite sequencing (RRBS). These new technologies did prompt the methylation studies, but also generated massive methylation sequencing data. Therefore, a huge challenge has emerged for researchers:how to quickly dig out the useful information from these massive data. With the hope of providing facilitations for researchers on DNA methylation, we developed the following analysis tools based on high-throughput sequencing, including simulation of methylation sequencing data, differentially methylated region (DMR) analysis, integrated platform for methylation analysis and visualization software.Firstly, we developed BSSim, a simulation software applied to generate short reads with sequencing format of main sequencing platforms. BSSim can evaluate the effects of multilevel factors on the sequencing data, also act as a reference for selecting the appropriate parameters in subsequent analysis and reduce the effect of error factors on the sequencing data.Secondly, we developed the swDMR, which is based on sliding window, to scan DMR at single-base resolution on the genome-wide scale. swDMR integrates a variety of commonly used statistical methods for detection, annotation and visualization of DMR from either paired samples or multiple samples treated by bisulfite. By using swDMR, users can identify potential functional regions involved in epigenetic regulation.Thirdly, we developed RRBS-Analyser:an integrated analysis platform which can comprehensively analyze RRBS sequencing data in genome-wide. It can perform quality assessment for the original sequencing data, produce basic statistical information, align the clean reads after quality control to the reference genome, identify and annotate sites of methylated cytosine, and identity and annotate DMR in multiple samples. Additionally, RRBS-Analyser supports genomic DNA methylation analysis of other species.Finally, we developed iMethy, a genome-wide methylation visualization software. It is characterized by the following functionalities:alignment, methylation identification and methylation visualization. It is powerful, userfriendly and can be used to effectively discover hidden features and patterns. Furthermore, iMethy is applicable to multiple operating systems, almost importantly, it has a fast rate in visualization refreshing and provide a powerful visualization tool for researchers on methylation analysis.