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Updated: Oct 15, 2025
Targeted DNA Methylation Analysis by Next-generation Sequencing
Published on: February 24, 2015
Repeatability of methylation measures using a QIAseq targeted methyl panel and comparison with the Illumina
Chenglong Yu1, Pierre-Antoine Dugué1,2,3, James G Dowty3
1Precision Medicine, School of Clinical Sciences at Monash Health, Monash University, Victoria, Australia.
Objective:
In previous studies using Illumina Infinium methylation arrays, we have identified DNA methylation marks associated with cancer predisposition and progression. In the present study, we have sought to find appropriate technology to both technically validate our data and expand our understanding of DNA methylation in these genomic regions. Here, we aimed to assess the repeatability of methylation measures made using QIAseq targeted methyl panel and to compare them with those obtained from the Illumina HumanMethylation450 (HM450K) assay. We included in the analysis high molecular weight DNA extracted from whole blood (WB) and DNA extracted from formalin-fixed paraffin-embedded tissues (FFPE).
Results:
The repeatability of QIAseq-methylation measures was assessed at 40 CpGs, using the Intraclass Correlation Coefficient (ICC). The mean ICCs and 95% confidence intervals (CI) were 0.72 (0.62-0.81), 0.59 (0.47-0.71) and 0.80 (0.73-0.88) for WB, FFPE and both sample types combined, respectively. For technical replicates measured using QIAseq and HM450K, the mean ICCs (95% CI) were 0.53 (0.39-0.68), 0.43 (0.31-0.56) and 0.70 (0.59-0.80), respectively. Bland-Altman plots indicated good agreement between QIAseq and HM450K measurements. These results demonstrate that the QIAseq targeted methyl panel produces reliable and reproducible methylation measurements across the 40 CpGs that were examined.
