Degradome sequencing provides large amounts of data regarding RNA degradation. The degradome library construction described here is modified from the 5'-rapid amplification of cDNA ends (5'-RACE), and each degradome cDNA is sequenced by next-generation sequencing (NGS). Degradome profiles provide information confirming miRNA-mediated cleavage of target genes and allow the identification of novel targets. Furthermore, degradome sequencing provides additional information for the study of RNA processing, such as information regarding RNA-binding proteins. In this chapter, we describe a detailed optimized protocol for constructing a degradome library with high yield and quality, along with NGS and data mining procedures. We hope that the degradome approach will be applied to further studies of non-model organisms.
Keywords: 5′-RACE; Degradome; Next-generation sequencing; RNA degradation; Target RNA; microRNA.