ArticleBio-protocol2023
Expression Stability Analysis of Candidate References for Normalization of RT-qPCR Data Using RefSeeker R package.
Article in Bio-protocol, 2023. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 6 papers.
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Who cites it
6 citing papers in PubMed.
- Identification and validation of stable reference genes for RT-qPCR analysis in blood samples from golden Syrian hamsters under different immunological conditions.Molecular biology reports · 2026Article
- Identification of condition-specific optimal reference genes for qPCR data normalisation in pink bollworm Pectinophora gossypiella (Saunders).Molecular biology reports · 2026Article
- The impact of copper status on lubabegron fumarate-induced growth and adipose immune cell infiltration in beef feedlot steers.Journal of animal science · 2026Article
- Identification and Validation of Reference Genes for Quantitative Real-Time PCR inPlants (Basel, Switzerland) · 2025Article
- microRNAs for qPCR Normalization Under Morphofunctional Conditions in Bovine Sperm (Bos taurus).Molecular reproduction and development · 2025Article
- Identifying the stability of housekeeping genes to be used for the quantitative real-time PCR normalization in retinal tissue of streptozotocin-induced diabetic rats.International journal of ophthalmology · 2024Article
Corrections and comments
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Authors and funding
4 authors.
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Abstract
When performing expression analysis either for coding RNA (e.g., mRNA) or non-coding RNA (e.g., miRNA), reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) is a widely used method. To normalize these data, one or more stable endogenous references must be identified. RefFinder is an online web-based tool using four almost universally used algorithms for assessing candidate endogenous references-delta-Ct, BestKeeper, geNorm, and Normfinder. However, the online interface is presently cumbersome and time consuming. We developed an R package, RefSeeker, which performs easy and straightforward RefFinder analysis by enabling raw data import and calculation of stability from each of the algorithms and provides data output tools to create graphs and tables. This protocol uses RefSeeker R package for fast and simple RefFinder stability analysis. Key features Perform stability analysis using five algorithms: Normfinder, geNorm, delta-Ct, BestKeeper, and RefFinder. Identification of endogenous references for normalization of RT-qPCR data. Create publication-ready graphs and tables output. Step-by-step guide dialog window for novice R users.
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