ArticleGenome biology2018
Comparison of whole-genome bisulfite sequencing library preparation strategies identifies sources of biases affecting DNA methylation data.
Article in Genome biology, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 176 papers, 2 of them syntheses that pooled it.
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Who cites it
176 citing papers in PubMed, 2 syntheses or guidelines pooled it.
- Epigenetic mechanisms in schizophrenia and other psychotic disorders: a systematic review of empirical human findings.Molecular psychiatry · 2020Pooled it
- Active Demethylation of Non-CpG Moieties in Animals: A Neglected Research Area.International journal of molecular sciences · 2019Pooled it
- Prediagnostic White Blood Cell DNA Methylation and Risk of Breast Cancer in the Prostate Lung, Colorectal, and Ovarian Cancer Screening Trial (PLCO) Cohort.Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology · 2021Trial
- Integrated pangenomics and population genomics reveal climate adaptation and maladaptation in an East Asian tree.Nature communications · 2026Article
- Long-read sequencing of single cell-derived melanoma sublines reveals divergent and parallel genomic and epigenomic evolutionary trajectories.Nature communications · 2026Article
- FINE-EM-seq: a rapid isothermal amplification method enabling comprehensive methylome profiling of zebrafish early embryos.Cell insight · 2026Article
- Maternal Sleep Quality and Early-Life Epigenetic Programming of Bone-Related Genes in Neonatal Cord Blood: Findings from the PERSIAN Birth Cohort, Isfahan.Calcified tissue international · 2026Article
- Circulating cell-free methylated DNA immunoprecipitation sequencing (cfMeDIP-seq) in oncology: from technological advances to clinical applications.NPJ precision oncology · 2026Review
- DNMT/TET Imbalance and Network-Level DNA Methylation Remodeling in Ovarian Aging: Mechanistic Perspectives.Biology · 2026Review
- The effects of ingested cellulose nanomaterials on DNA methylation in intestinal cells.Particle and fibre toxicology · 2026Article
- Cancer-Like Fragmentomic Characteristics of Somatic Variants in Cell-Free DNA.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2026Article
- Naturally-occurring DNA fragment termini correlate with methylation at CpG sites in hair and blood plasma cell-free DNA.BMC genomics · 2026Article
- Dark and camouflaged genomic regions remain challenging in CHM13.Scientific reports · 2026Article
- m6AConquer: a consistently quantified and orthogonally validated database for the N6-methyladenosine (m6A) epitranscriptome.Nucleic acids research · 2026Article
- 5mC and 5hmC methylation sequencing: the power of 6-base sequencing in a multiomic era.Epigenomics · 2026Review
- Application of a modified MSRE-qPCR method for detecting circulating cell-free DNA methylation in cervical cancer.Frontiers in oncology · 2026Article
- Circulating neuron-derived cfDNA for blood-based detection of Alzheimer's and other neurodegenerative conditions.Frontiers in neurology · 2026Article
- Laser Capture Microdissection Followed by DNA Methylation Profiling.Methods in molecular biology (Clifton, N.J.) · 2026Article
- The INSPECTOR study: enhanced feasibility for clinical translation of a multi-cancer early detection method based on enzyme-assisted high signal-to-noise ratio sequencing of methylated circulating tumor DNA.Cancer communications (London, England) · 2025Article
- From population science to the clinic? Limits of epigenetic clocks as personal biomarkers.Epigenomics · 2025Article
116 more citing papers are in PubMed but not listed here.
Corrections and comments
- Erratum issued
Authors and funding
7 authors.
Funding
Abstract
backgroundWhole-genome bisulfite sequencing (WGBS) is becoming an increasingly accessible technique, used widely for both fundamental and disease-oriented research. Library preparation methods benefit from a variety of available kits, polymerases and bisulfite conversion protocols. Although some steps in the procedure, such as PCR amplification, are known to introduce biases, a systematic evaluation of biases in WGBS strategies is missing.
resultsWe perform a comparative analysis of several commonly used pre- and post-bisulfite WGBS library preparation protocols for their performance and quality of sequencing outputs. Our results show that bisulfite conversion per se is the main trigger of pronounced sequencing biases, and PCR amplification builds on these underlying artefacts. The majority of standard library preparation methods yield a significantly biased sequence output and overestimate global methylation. Importantly, both absolute and relative methylation levels at specific genomic regions vary substantially between methods, with clear implications for DNA methylation studies.
conclusionsWe show that amplification-free library preparation is the least biased approach for WGBS. In protocols with amplification, the choice of bisulfite conversion protocol or polymerase can significantly minimize artefacts. To aid with the quality assessment of existing WGBS datasets, we have integrated a bias diagnostic tool in the Bismark package and offer several approaches for consideration during the preparation and analysis of WGBS datasets.
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