MBDDiff: an R package designed specifically for processing MBDcap-seq datasets

Yuanhang Liu, Desiree Wilson, Robin J. Leach, Yidong Chen

Since its initial discovery in 1975, DNA methylation has been intensively studied and shown to be involved in various biological processes, such as development, aging and tumor progression. Many experimental techniques have been developed to measure the level of DNA methylation. Methyl-CpG binding domain-based capture followed by high-throughput sequencing (MBDCap-seq) is a widely used method for characterizing DNA methylation patterns in a genome-wide manner. However, current methods for processing MBDCap-seq datasets does not take into account of the region-specific genomic characteristics that might have an impact on the measurements of the amount of methylated DNA (signal) and background fluctuation (noise). Thus, specific software needs to be developed for MBDCap-seq experiments. A new differential methylation quantification algorithm for MBDCap-seq, MBDDiff, was implemented. To evaluate the performance of the MBDDiff algorithm, a set of simulated signal based on negative binomial and Poisson distribution with parameters estimated from real MBDCap-seq datasets accompanied with different background noises were generated, and then performed against a set of commonly used algorithms for MBDCap-seq data analysis in terms of area under the ROC curve (AUC), number of false discoveries and statistical power. In addition, we also demonstrated the effective of MBDDiff algorithm to a set of in-house prostate cancer samples, endometrial cancer samples published earlier, and a set of public-domain triple negative breast cancer samples to identify potential factors that contribute to cancer development and recurrence. In this paper we developed an algorithm, MBDDiff, designed specifically for datasets derived from MBDCap-seq. MBDDiff contains three modules: quality assessment of datasets and quantification of DNA methylation; determination of differential methylation of promoter regions; and visualization functionalities. Simulation results suggest that MBDDiff performs better compared to MEDIPS and DESeq in terms of AUC and the number of false discoveries at different levels of background noise. MBDDiff outperforms MEDIPS with increased backgrounds noise, but comparable performance when noise level is lower. By applying MBDDiff to several MBDCap-seq datasets, we were able to identify potential targets that contribute to the corresponding biological processes. Taken together, MBDDiff provides user an accurate differential methylation analysis for data generated by MBDCap-seq, especially under noisy conditions.