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Comparison of nanopore sequencing, MethylationEPIC array, and EM-Seq for DNA methylation detection

Year: 2026

Type: article

Abstract: DNA methylation is an important biological process in epigenetics, and many methods have been developed to profile DNA methylation. Recent studies have employed Oxford Nanopore long-read sequencing for DNA methylation detection, presenting an alternative to the widely utilized Infinium arrays and short-read methods. In this study, we evaluate the performance of Nanopore sequencing in DNA methylation detection by comparing it to Illumina's MethylationEPIC microarray (EPIC) and Enzymatic Methyl-Sequencing (EM-Seq). The initial comparison was between Nanopore and the EPIC array using blood samples (n = 4). Among the ∼850,000 CpG sites covered by both methods, we observed high concordance (Pearson correlation coefficient, r ≥ 0.94 across all four samples). After downsampling Nanopore data from an average coverage of 26.4 reads per site to 10 reads per site, the correlation in CpG methylation remained high (r ≥ 0.93). When comparing Nanopore and EM-Seq using brain tissue samples (n = 4), lower correlation of CpG methylation (r: 0.79-0.88) was detected between Nanopore and EM-Seq, which can be attributed to biased and reduced coverage of hypomethylated CpG sites by EM-Seq. We also investigated additional features of Nanopore sequencing, such as native DNA sequencing that can differentiate 5mC and 5hmC, as well as haplotype phasing. Overall, the Nanopore platform exhibited a high degree of concordance with the EPIC array and provided more uniform genomic coverage than EM-Seq. This study provides insights for researchers in selecting appropriate DNA methylation detection methods, considering factors such as cost, DNA input, and the complexity of downstream analysis.

Source: Computational Biology and Chemistry

DOI: 10.1016/j.compbiolchem.2026.109204