A scaling normalization method for differential expression analysis of RNA-seq data
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The fine detail provided by sequencing-based transcriptome surveys suggests that RNA-seq is likely to become the platform of choice for interrogating steady state RNA. In order to discover biologically important changes in expression, we show that normalization continues to be an essential step in the analysis. We outline a simple and effective method for performing normalization and show dramatically improved results for inferring differential expression in simulated and publicly available data sets.
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Normalization methods TMM, DESeq, PoissonSeq, and UpperQuartile take heterogeneous transcript distributions into account by ignoring highly variable and/or highly expressed features.
“Normalization methods that take this into account are TMM [47], DESeq [48], PoissonSeq [49] and UpperQuartile [45], which ignore highly variable and/or highly expressed features.”
From A survey of best practices for RNA-seq data analysis (2016) · cites this paper as [47]
voom has the ability to respect normalization factors computed outside of limma by methods such as trimmed mean of M-values or conditional quantile normalization.
“Alternatively, voom has the ability to respect normalization factors computed outside of limma by methods such as trimmed mean of M-values (41) or conditional quantile normalization (42).”
From limma powers differential expression analyses for RNA-sequencing and microarray studies (2015) · cites this paper as [41]
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- Authors
- Robinson, M. D., & Oshlack, A.
- Affiliations
- Garvan Institute of Medical Research (AU)
- Walter and Eliza Hall Institute of Medical Research (AU)
- Corresponding author
- Robinson, Mark D
- Oshlack, Alicia
- Venue
- Genome Biology
- Published
- 2010-03-02
- Volume
- 11
- Issue
- 3
- Pages
- R25-R25
- ISSN
- 1465-6906 (ISSN-L)
- 1474-760X
- PMC
- PMC2864565
- OpenAlex
- W2107018762
- Cited by
- 7,876
- References
- 25
- Citations by year
- 2026: 452 · 2025: 604 · 2024: 832 · 2023: 700 · 2022: 773 · 2021: 1010 · 2020: 730 · 2019: 610 · 2018: 540 · 2017: 440 · 2016: 348 · 2015: 285 · 2014: 222 · 2013: 153 · 2012: 100
- FWCI
- 41.77
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Locations
- Genome biology (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Minerva Access (University of Melbourne) (submittedVersion, cc-by)
- Funders
- Medical Research Council
- National Health and Medical Research Council (490037)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Gene expression and cancer classification
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
- Keywords
- RNA-seq, normalization, differential gene expression, transcriptome, gene expression, read count normalization
- MeSH terms
- Base Sequence, Computer Simulation, RNA, Models, Statistical, Gene Library, Gene Expression Profiling
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