Comparison and imputation-aided integration of five commercial platforms for targeted DNA methylome analysis

Targeted bisulfite sequencing (TBS) has become the method of choice for the cost-effective, targeted analysis of the human methylome at base-pair resolution. In this study, we benchmarked five commercially available TBS platforms-three hybridization capture-based (Agilent, Roche and Illumina) and two reduced-representation-based (Diagenode and NuGen)-across 11 samples. Two samples were also compared with whole-genome DNA methylation sequencing with the Illumina and Oxford Nanopore platforms. We assessed workflow complexity, on/off-target performance, coverage, accuracy and reproducibility. Although all platforms produced robust and reproducible data, major differences in the number and identity of the CpG sites covered make it difficult to compare datasets generated on different platforms. To overcome this limitation, we applied imputation and show that it improves interoperability from an average of 10.35% (0.8 million) to 97% (7.6 million) common CpG sites. Our study provides guidance on which TBS platform to use for different methylome features and offers an imputation-based harmonization solution that allows comparative, integrative analysis. Differences in CpG coverage of targeted bisulfite sequencing methods can be overcome using imputation.

Automated summary

Targeted bisulfite sequencing (TBS) is a cost-effective method for analyzing the human methylome. In this study, five commercially available TBS platforms were compared. The results showed that although all platforms produced robust and reproducible data, there were major differences in the CpG sites covered. However, these differences can be overcome using imputation. The study provides guidance on choosing TBS platforms and offers a harmonization solution for comparative analysis.