Degradome Sequencing in Plants
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.
Key wordsDegradome RNA degradation microRNA Target RNA 5′-RACE Next-generation sequencing
We are grateful to the High Throughput Genomics Core at the Biodiversity Research Center, Academia Sinica, for performing Illumina sequencing. The study described in this chapter was supported by a grant from the Ministry of Science and Technology of Taiwan under contract number MOST 106-2321-B-002-008.
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