**RNA‐seq: Applications and Best Practices**

**RNA**‐**seq: Applications and Best Practices**

DOI: 10.5772/intechopen.69250

#### Michele Araújo Pereira, Eddie Luidy Imada and Rafael Lucas Muniz Guedes and Rafael Lucas Muniz Guedes Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

Michele Araújo Pereira, Eddie Luidy Imada

http://dx.doi.org/10.5772/intechopen.69250

#### **Abstract**

RNA‐sequencing (RNA‐seq) is the state‐of‐the‐art technique for transcriptome analysis that takes advantage of high‐throughput next‐generation sequencing. Although being a powerful approach, RNA‐seq imposes major challenges throughout its steps with numerous caveats. There are currently many experimental options available, and a com‐ plete comprehension of each step is critical to make right decisions and avoid getting into inconclusive results. A complete workflow consists of: (1) experimental design; (2) sample and library preparation; (3) sequencing; and (4) data analysis. RNA‐seq enables a wide range of applications such as the discovery of novel genes, gene/transcript quan‐ tification, and differential expression and functional analysis. This chapter will encom‐ pass the main aspects from sample preparation to downstream data analysis. It will be discussed how to obtain high‐quality samples, replicates amount, library preparation, sequencing platforms and coverage, focusing on best recommended practices based on specialized literature. Basic techniques and well‐known algorithms are presented and discussed, guiding both beginners and experienced users in the implementation of reli‐ able experiments.

**Keywords:** RNA‐seq, next‐generation sequencing, transcriptome, data analysis, best practices
