ArticleFrontiers in oncology2019
Standardization of Sequencing Coverage Depth in NGS: Recommendation for Detection of Clonal and Subclonal Mutations in Cancer Diagnostics.
Article in Frontiers in oncology, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 86 papers, 2 of them syntheses that pooled it.
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Who cites it
86 citing papers in PubMed, 2 syntheses or guidelines pooled it.
- Comparison of liquid-based to tissue-based biopsy analysis by targeted next generation sequencing in advanced non-small cell lung cancer: a comprehensive systematic review.Journal of cancer research and clinical oncology · 2020Pooled it
- Resistance-Associated Mutations in Chronic Lymphocytic Leukemia Patients Treated With Novel Agents.Frontiers in oncology · 2020Pooled it
- Exome sequencing shows same pattern of clonal tumor mutational burden, intratumor heterogenicity and clonal neoantigen between autologous tumor and Vigil product.Scientific reports · 2025Trial
- The HBV basal core promoter mutation confers a replicative advantage and transcriptionally reprograms hepatocytes toward HCC subtypes.bioRxiv : the preprint server for biology · 2026Article
- Circulating tumor DNA-based detection of molecular residual disease in isocitrate dehydrogenase-mutant cholangiocarcinoma for biomarker-guided therapy.Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico · 2026Review
- Landscape of parental postzygotic mutations across >11,000 rare disease trios.American journal of human genetics · 2026Article
- Human adipose-derived mesenchymal stromal cells in vitro expansion for regenerative applications: tracking small genetic variants reveals dynamic changes within a genetically stable framework.Stem cell research & therapy · 2026Article
- Somatic mutation profiling by NGS-based liquid biopsy in advanced non-small cell lung cancer: frequency, clinical correlates, and prognostic significance.Molecular biology reports · 2026Article
- Expert recommendations for PIK3CA testing in HR+/HER2- locally advanced and metastatic breast cancer.NPJ breast cancer · 2026Review
- Article
- Exploring alterations in the gut resistome in medically treated inflammatory bowel disease patients.BMC microbiology · 2026Article
- Advancing the Diagnosis of Non-Hodgkin Lymphoma Through Next-Generation Sequencing in Developing Countries: An Evaluation of Progress-A Narrative Review.Health science reports · 2026Article
- SARS-CoV-2 Error Catastrophe Under Molnupiravir: Mutagenic Enhancement Enables Viral Persistence with Impaired Fitness.Viruses · 2026Article
- Emerging role of low-frequency somatic mutations in cancer relapse: from early detection to precision oncology.Frontiers in oncology · 2026Review
- Unmasking the "targetless" illusion: branched clonal evolution ofFrontiers in immunology · 2026Article
- How to Read a Next-Generation Sequencing Report for AML and MDS? What Hematologists Need to Know.Journal of clinical medicine · 2025Review
- Evaluation of false positive and false negative errors in targeted next generation sequencing.Genome biology · 2025Article
- Functional genomics and tumor microenvironment analysis reveal prognostic biological subtypes in Mantle cell lymphoma.Nature communications · 2025Article
- Characterization of the mutational status of glioblastoma and high-grade astrocytomas in a Latin American cohort.Scientific reports · 2025Article
- Variation of and associations with the depth and evenness of sequencing coverage in archived plastid genomes.Scientific reports · 2025Article
26 more citing papers are in PubMed but not listed here.
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Authors and funding
8 authors.
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Abstract
The insufficient standardization of diagnostic next-generation sequencing (NGS) still limits its implementation in clinical practice, with the correct detection of mutations at low variant allele frequencies (VAF) facing particular challenges. We address here the standardization of sequencing coverage depth in order to minimize the probability of false positive and false negative results, the latter being underestimated in clinical NGS. There is currently no consensus on the minimum coverage depth, and so each laboratory has to set its own parameters. To assist laboratories with the determination of the minimum coverage parameters, we provide here a user-friendly coverage calculator. Using the sequencing error only, we recommend a minimum depth of coverage of 1,650 together with a threshold of at least 30 mutated reads for a targeted NGS mutation analysis of ≥3% VAF, based on the binomial probability distribution. Moreover, our calculator also allows adding assay-specific errors occurring during DNA processing and library preparation, thus calculating with an overall error of a specific NGS assay. The estimation of correct coverage depth is recommended as a starting point when assessing thresholds of NGS assay. Our study also points to the need for guidance regarding the minimum technical requirements, which based on our experience should include the limit of detection (LOD), overall NGS assay error, input, source and quality of DNA, coverage depth, number of variant supporting reads, and total number of target reads covering variant region. Further studies are needed to define the minimum technical requirements and its reporting in diagnostic NGS.
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