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Updated: Jun 24, 2026
Targeted DNA Methylation Analysis by Next-generation Sequencing
Published on: February 24, 2015
Rapid and quantitative detection of CpG-methylation status using TaqMan PCR combined with methyl-binding-domain
Naoki Kimura1, Toyoki Moribe, Norio Iizuka
1Research Group, Molecular Diagnostics R&D Department, Roche Diagnostics K.K., 6-1, Shiba 2-chome, Minato-ku, Tokyo 105-0014, Japan.
Objectives:
To assess the medical applicability of CpG methylation as molecular markers for cancer diagnosis, we established a new system to determine DNA methylation based on TaqMan PCR combined with a methyl-binding-domain polypeptide 2.
Design And Methods:
We evaluated the diagnostic applicability of this approach by examining the methylation status of two tumor suppressor genes, RASSF1A and APC, in 10 paired hepatocellular carcinoma (HCC) and the corresponding non-tumor liver tissues.
Results:
Methylation levels of total 20 clinical samples measured by the TaqMan PCR assay showed a significantly positive correlation (R=0.814, P<0.0005 for RASSF1A, R=0.736, P<0.00001 for APC) with those calculated by bisulfite sequencing. The methylated DNA amount measured by our TaqMan PCR system precisely replicated the methylation status estimated by direct sequencing.
Conclusions:
This suggests our method may serve as a reliable and easy-to-use tool for cancer diagnosis using methylated genes as biomarkers.
