ArticleHuman mutation2020
Quantification of DNA methylation independent of sodium bisulfite conversion using methylation-sensitive restriction enzymes and digital PCR.
Article in Human mutation, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed.
- Recent advances on gene-related DNA methylation in cancer diagnosis, prognosis, and treatment: a clinical perspective.Clinical epigenetics · 2025Review
- Digital PCR-based genetic profiling from vitreous fluid as liquid biopsy for primary uveal melanoma: a proof-of-concept study.Journal of experimental & clinical cancer research : CR · 2025Article
- Review
- Methylation‑sensitive restriction enzyme‑droplet digital PCR assay for the one‑step highly sensitive analysis of DNA methylation hotspots.International journal of molecular medicine · 2024Article
- Investigating FGFR2 gene as a blood-based epigenetic biomarker in gastric cancer.Molecular biology reports · 2024Article
- Density Control over MBD2 Receptor-Coated Surfaces Provides Superselective Binding of Hypermethylated DNA.ACS applied materials & interfaces · 2022Article
- RASSF1A methylation as a biomarker for detection of colorectal cancer and hepatocellular carcinoma.World journal of gastrointestinal oncology · 2022Article
- Quantification of Unmethylated Insulin DNA Using Methylation Sensitive Restriction Enzyme Digital Polymerase Chain Reaction.Transplant international : official journal of the European Society for Organ Transplantation · 2022Article
- Quantification of DNA methylation independent of sodium bisulfite conversion using methylation-sensitive restriction enzymes and digital PCR.Human mutation · 2020Article
- Interplay between TERT promoter mutations and methylation culminates in chromatin accessibility and TERT expression.PloS one · 2020Article
Corrections and comments
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Authors and funding
6 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Epigenetic regulation is important in human health and disease, but the exact mechanisms remain largely enigmatic. DNA methylation represents one epigenetic aspect but is challenging to quantify. In this study, we introduce a digital approach for the quantification of the amount and density of DNA methylation. We designed an experimental setup combining efficient methylation-sensitive restriction enzymes with digital polymerase chain reaction (PCR) to quantify a targeted density of DNA methylation independent of bisulfite conversion. By using a stable reference and comparing experiments treated and untreated with these enzymes, copy number instability could be properly normalized. In silico simulations demonstrated the mathematical validity of the setup and showed that the measurement precision depends on the amount of input DNA and the fraction methylated alleles. This uncertainty could be successfully estimated by the confidence intervals. Quantification of RASSF1 promoter methylation in a variety of healthy and malignant samples and in a calibration curve confirmed the high accuracy of our approach, even in minute amounts of DNA. Overall, our results indicate the possibility of quantifying DNA methylation with digital PCR, independent of bisulfite conversion. Moreover, as the context-density of methylation can also be determined, biological mechanisms can now be quantitatively assessed.
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