Performance comparison and in-silico harmonisation of commercial platforms for DNA methylome analysis by targeted bisulfite sequencing

Abstract

DNA methylation is a key epigenetic modification in the regulation of cell fate and differentiation, and its analysis is gaining increasing importance in both basic and clinical research. Targeted Bisulfite Sequencing (TBS) has become the method of choice for the cost-effective, targeted analysis of the human methylome at base-pair resolution. Here we benchmarked five commercially available TBS platforms, including three hybridization capture-based (Agilent, Roche, and Illumina) and two RRBS-based (Diagenode and NuGen), across 16 samples. A subset of these were also compared to whole-genome DNA methylation sequencing with the Illumina and Oxford Nanopore platforms. We assessed performance with respect to workflow complexity, on/off-target performance, coverage, accuracy and reproducibility. We find all platforms able to produce usable data but major differences for some performance criteria, especially in the number and identity of the CpG sites covered, which affects the interoperability of datasets generated on these different platforms. To overcome this limitation, we used imputation and show that it improves the interoperability from an average of 10.35% (0.8M CpG sites) to 97% (7.6M CpG sites). Our study provides cross-validated guidance on which TBS platform to use for different features of the methylome and offers an imputation-based harmonization solution for improved interoperability between platforms, allowing comparative and integrative analysis.

A Comparative Overview of Epigenomic Profiling Methods

“…For the development of reliable best practices guidelines, direct and side-by-side comparisons of epigenomic assays, such as previous comparative studies (Harris et al, 2010;Chang et al, 2018;Kacmarczyk et al, 2018;Gontarz et al, 2020;Heiss et al, 2020;Tanić et al, 2021) are required. Additional analyses are called for to clearly elucidate advantages and disadvantages of each of the methods in a comprehensive manner, perhaps in community efforts in the future.…”

“…TruSeq EPIC has a significant improvement over EPIC-array regarding genomic resolution and number of CpGs, however it suffers from lower precision due to limited coverage per site for comparable costs with arrays (Heiss et al, 2020). Other commercially available targeted bisulfite sequencing platforms also exist with comparable characteristics with the TruSeq (Tanić et al, 2021). The significant limitations of methods based on bisulfite conversion remain cost, incomplete conversion, as well as partial DNA digestion during bisulfite treatment.…”

“…It has thus become increasingly critical to choose the right type of assay depending on the goals, context, and applications in mind. Recently, the number of comparative studies using experimental or simulated data for DNA methylome (Kacmarczyk et al, 2018;Heiss et al, 2020;Tanić et al, 2021) and chromatin accessibility assays (Chang et al, 2018;Gontarz et al, 2020) has increased. However, clear guidelines that integrate the available information to help researchers base their choice among the large selection of epigenomic assays on the most relevant criteria are lacking.…”