Assembly of RNA ‐Seq Short Reads into Transcriptome Sequences

RNA-Seq is a technology to sequence transcriptomes using next-generation sequencing technologies. It has been widely used for analyses such as gene expression profiling and identification of differentially expressed genes (DEG). This chapter focuses on the design of RNA-Seq experiments and on the bioinformatics issues related to the assembly of RNA-Seq short reads into reference transcriptomes. It presents procedures and command lines for both de novo assembly approaches and reference-sequence-guided assembly approaches. In spite of the rapid progress in genome sequencing with aquaculture species, the reference genome sequences or reference transcriptomes are not yet available formost aquaculture species. If a reference genome sequence is available, reference-guided assembly methods can be used. In contrast, de novo RNA-Seq assembly methods must be used in the absence of a reference genome sequence. TopHat-Cufflinks is the most popular reference-guided assembly method, while Trinity is the most popular de novo assembly method.