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Statistical Genomics pp 391–416Cite as

It’s DE-licious: A Recipe for Differential Expression Analyses of RNA-seq Experiments Using Quasi-Likelihood Methods in edgeR

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1418))

Abstract

RNA sequencing (RNA-seq) is widely used to profile transcriptional activity in biological systems. Here we present an analysis pipeline for differential expression analysis of RNA-seq experiments using the Rsubread and edgeR software packages. The basic pipeline includes read alignment and counting, filtering and normalization, modelling of biological variability and hypothesis testing. For hypothesis testing, we describe particularly the quasi-likelihood features of edgeR. Some more advanced downstream analysis steps are also covered, including complex comparisons, gene ontology enrichment analyses and gene set testing. The code required to run each step is described, along with an outline of the underlying theory. The chapter includes a case study in which the pipeline is used to study the expression profiles of mammary gland cells in virgin, pregnant and lactating mice.

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Acknowledgements

This worked was funded by the University of Melbourne (Elizabeth and Vernon Puzey Scholarship to Aaron T.L. Lun), by the National Health and Medical Research Council (NHMRC) (Fellowship 1058892 and Program 1054618 to Gordon K. Smyth), by the NHMRC Independent Research Institutes Infrastructure Support (IRIIS) Scheme, and by a Victorian State Government Operational Infrastructure Support (OIS) Grant.

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Authors and Affiliations

  1. Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia

    Aaron T. L. Lun, Yunshun Chen & Gordon K. Smyth

Authors
  1. Aaron T. L. Lun
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  2. Yunshun Chen
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  3. Gordon K. Smyth
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Editor information

Editors and Affiliations

  1. Ohio State University, Biomed Informatics, College of Medicine, Columbus, Ohio, USA

    Ewy Mathé

  2. National Cancer Institute, National Institutes of Health, Columbia, Maryland, USA

    Sean Davis

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Lun, A.T.L., Chen, Y., Smyth, G.K. (2016). It’s DE-licious: A Recipe for Differential Expression Analyses of RNA-seq Experiments Using Quasi-Likelihood Methods in edgeR. In: Mathé, E., Davis, S. (eds) Statistical Genomics. Methods in Molecular Biology, vol 1418. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3578-9_19

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  • DOI: https://doi.org/10.1007/978-1-4939-3578-9_19

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3576-5

  • Online ISBN: 978-1-4939-3578-9

  • eBook Packages: Springer Protocols

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