Talking the talk, but not walking the walk: RT-qPCR as a paradigm for the lack of reproducibility in molecular research
Open accessAbstract
Poorly executed and inadequately reported molecular measurement methods are amongst the causes underlying the lack of reproducibility of much biomedical research. Although several high impact factor journals have acknowledged their past failure to scrutinise adequately the technical soundness of manuscripts, there is a perplexing reluctance to implement basic corrective measures. The reverse transcription real-time quantitative PCR (RT-qPCR) is probably the most straightforward measurement technique available for RNA quantification and is widely used in research, diagnostic, forensic and biotechnology applications. Despite the impact of the minimum information for the publication of quantitative PCR experiments (MIQE) guidelines, which aim to improve the robustness and the transparency of reporting of RT-qPCR data, we demonstrate that elementary protocol errors, inappropriate data analysis and inadequate reporting continue to be rife and conclude that the majority of published RT-qPCR data are likely to represent technical noise.
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Independent studies have reported that the identities of some nucleotide sequence reagents do not match their descriptions.
“Independent studies have described examples where the identities of some nucleotide sequence reagents do not match their descriptions (5–17), including oligonucleotide probes on microarrays (18, 19), and shRNAs in libraries (20).”
From Misspellings or “miscellings”—Non‐verifiable and unknown cell lines in cancer research publications (2024) · cites this paper as [12]
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Studies on Scratch Tests in Atopic Dermatitis, and Comparison with the Intracutaneous Method*†
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Flagged first, then by citations
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miRNA extracted from extracellular vesicles is a robust biomarker of amyotrophic lateral sclerosis
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Identification of human gene research articles with wrongly identified nucleotide sequences
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Misspellings or “miscellings”—Non‐verifiable and unknown cell lines in cancer research publications
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- Authors
- Bustin, S., & Nolan, T.
- Affiliations
- Anglia Ruskin University (GB)
- University of Manchester (GB)
- Corresponding author
- Bustin, Stephen
- ORCID
- Bustin, Stephen
- Venue
- European Journal of Clinical Investigation
- Publisher
- Wiley
- Published
- 2017-09-02
- Volume
- 47
- Issue
- 10
- Pages
- 756-774
- ISSN
- 0014-2972 (ISSN-L)
- OpenAlex
- W2743105407
- Cited by
- 113
- References
- 165
- Citations by year
- 2026: 7 · 2025: 15 · 2024: 14 · 2023: 8 · 2022: 16 · 2021: 13 · 2020: 14 · 2019: 17 · 2018: 8 · 2017: 1
- FWCI
- 1.02
- Type
- journal article
- Language
- en
- Open access
- green
- figshare.com
- Licence
- Locations
- European Journal of Clinical Investigation (publishedVersion)
- PubMed (publishedVersion)
- Anglia Ruskin Research Online (Anglia Ruskin University) (submittedVersion)
- Figshare (submittedVersion, cc-by)
- Research Explorer (The University of Manchester) (submittedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Molecular Biology Techniques and Applications
- Social Sciences › Decision Sciences › Statistics, Probability and Uncertainty › Meta-analysis and systematic reviews
- Health Sciences › Medicine › Physiology › Clinical Laboratory Practices and Quality Control
- Keywords
- RT-qPCR, reproducibility, MIQE guidelines, reporting quality, technical noise, RNA quantification, data analysis, research integrity
- MeSH terms
- Humans, RNA, Sensitivity and Specificity, Reproducibility of Results, Biomedical Research, Journal Impact Factor, Real-Time Polymerase Chain Reaction
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
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