ArticleScientific reports2016
Experimental factors affecting the robustness of DNA methylation analysis.
Article in Scientific reports, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers, 2 of them syntheses that pooled it.
What it found
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The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
13 citing papers in PubMed, 2 syntheses or guidelines pooled it, 21 citations in OpenAlex.
- Fecal DNA methylation markers for detecting stages of colorectal cancer and its precursors: a systematic review.Clinical epigenetics · 2020Pooled it
- Prognostic DNA methylation markers for sporadic colorectal cancer: a systematic review.Clinical epigenetics · 2018Pooled it
- DNA Methylation Analyses of Tissue Samples and Cell-Free DNA by Droplet Digital PCR.Methods in molecular biology (Clifton, N.J.) · 2026Article
- Article
- Review
- Promoter hypermethylation of RARB and GSTP1 genes in plasma cell-free DNA as breast cancer biomarkers in Peruvian women.Molecular genetics & genomic medicine · 2023Article
- Application of droplet digital polymerase chain reaction of plasma methylated septin 9 on detection and early monitoring of colorectal cancer.Scientific reports · 2021Article
- Multiregional assessment of CIMP in primary colorectal cancers: Phenotype concordance but marker variability.International journal of cancer · 2021Article
- Epigenetic Landscape of Liquid Biopsy in Colorectal Cancer.Frontiers in cell and developmental biology · 2021Review
- Comparison of Bisulfite Pyrosequencing and Methylation-Specific qPCR for Methylation Assessment.International journal of molecular sciences · 2020Article
- DNA-Methylation-Based Detection of Urological Cancer in Urine: Overview of Biomarkers and Considerations on Biomarker Design, Source of DNA, and Detection Technologies.International journal of molecular sciences · 2019Review
- A robust internal control for high-precision DNA methylation analyses by droplet digital PCR.Clinical epigenetics · 2018Article
- Prognostic relevance of an epigenetic biomarker panel in sentinel lymph nodes from colon cancer patients.Clinical epigenetics · 2017Article
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
9 authors at 3 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Diverging methylation frequencies are often reported for the same locus in the same disease, underscoring the need for limiting technical variability in DNA methylation analyses. We have investigated seven likely sources of variability at different steps of bisulfite PCR-based DNA methylation analyses using a fully automated quantitative methylation-specific PCR setup of six gene promoters across 20 colon cancer cell lines. Based on >15,000 individual PCRs, all tested parameters affected the normalized percent of methylated reference (PMR) differences, with a fourfold varying magnitude. Additionally, large variations were observed across the six genes analyzed. The highest variation was seen using single-copy genes as reference for normalization, followed by different amounts of template in the PCR, different amounts of DNA in the bisulfite reaction, and storage of bisulfite converted samples. Finally, when a highly standardized pipeline was repeated, the difference in PMR value for the same assay in the same cell line was on average limited to five (on a 0-100 scale). In conclusion, a standardized pipeline is essential for consistent methylation results, where parameters are kept constant for all samples. Nevertheless, a certain level of variation in methylation values must be expected, underscoring the need for careful interpretation of data.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.