Abstract
With the ability to obtain several millions of reads per sample, high-throughput RNA sequencing (RNA-Seq) enables investigation of any transcriptome at a fine resolution. Not just the messenger RNA (mRNA), but a wide variety of different RNA populations (e.g., total RNA, microRNA, long ncRNA, pre-mRNA) can also be investigated using RNA-Seq. While facilitating accurate quantification of gene expression, RNA-Seq offers the opportunity to estimate abundance of isoforms and find novel transcripts and allele-specific transcripts. In this chapter, we describe a protocol to construct an RNA-Seq library for sequencing on Illumina NGS platforms and a computational pipeline to perform RNA-Seq data analysis. The protocols described in this chapter can be applied to the analysis of differential gene expression in control versus 17β-estradiol treatment of in vivo or in vitro systems.
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Acknowledgments
We thank Dr. Thomas Girke at the University of California Riverside for sharing his R scripts.
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Withanage, M.H.H., Liang, H., Zeng, E. (2022). RNA-Seq Experiment and Data Analysis. In: Eyster, K.M. (eds) Estrogen Receptors. Methods in Molecular Biology, vol 2418. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1920-9_22
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DOI: https://doi.org/10.1007/978-1-0716-1920-9_22
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