Sequences to Differences in Gene Expression: Analysis of RNA-Seq Data.

RNA-Seq is now a routinely employed assay to measure gene expression. As the technique matured over the last decade, so have dedicated analytic tools. In this chapter, we first describe the mainstream as well as the most up-to-date protocols and their implications on downstream analysis. We then detail the steps entailing RNA-Seq analysis in three main stages: (i) preprocessing and data preparation, (ii) upstream processing, and (iii) high-level analyses. We review the most recent and relevant tools as one workflow following a stepwise order. The chapter further encompasses in-depth features of these tools. Details of the required code are made available throughout the chapter, as well as of the underlying statistics. We illustrate these steps with analysis of publicly available RNA-Seq data.