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

Area of Science:

• Epigenetics
• Genomics
• Molecular Biology

Background:

• DNA methylation is a key epigenetic mechanism regulating gene expression.
• Traditional methods like Infinium arrays and short-read sequencing are widely used for DNA methylation profiling.
• Oxford Nanopore long-read sequencing emerges as a novel alternative for DNA methylation detection.

Purpose of the Study:

• To evaluate the performance of Oxford Nanopore sequencing for DNA methylation detection.
• To compare Nanopore sequencing with Illumina MethylationEPIC array (EPIC) and Enzymatic Methyl-Sequencing (EM-Seq).
• To provide insights for selecting appropriate DNA methylation detection methods.

Main Methods:

• Comparison of Nanopore sequencing with EPIC array using blood samples (n=4).
• Comparison of Nanopore sequencing with EM-Seq using brain tissue samples (n=4).
• Analysis of CpG site concordance, genomic coverage, and additional Nanopore features (5mC/5hmC differentiation, haplotype phasing).

Main Results:

• High concordance (r ≥ 0.94) observed between Nanopore and EPIC array, even with downsampled Nanopore data (r ≥ 0.93).
• Lower correlation (r: 0.79-0.88) between Nanopore and EM-Seq, attributed to EM-Seq's biased coverage of hypomethylated sites.
• Nanopore sequencing demonstrated more uniform genomic coverage compared to EM-Seq.

Conclusions:

• Nanopore sequencing is a reliable method for DNA methylation detection, showing high concordance with established array technology.
• Nanopore offers advantages in uniform coverage and native DNA analysis capabilities (5mC/5hmC, haplotype phasing).
• Findings assist researchers in choosing DNA methylation profiling methods based on cost, input DNA, and analysis complexity.