Lin Shih-Shun, Chen Yihua, Lu Mei-Yeh Jade
Institute of Biotechnology, National Taiwan University, Taipei, Taiwan.
High Throughput Genomics Core, Biodiversity Research Center, Academia Sinica, Taipei, Taiwan.
Methods Mol Biol. 2019;1932:197-213. doi: 10.1007/978-1-4939-9042-9_15.
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.
降解组测序可提供大量有关RNA降解的数据。此处描述的降解组文库构建方法是对5'-cDNA末端快速扩增（5'-RACE）进行的改良，每个降解组cDNA通过新一代测序（NGS）进行测序。降解组图谱提供的信息可证实miRNA介导的靶基因切割，并有助于鉴定新的靶标。此外，降解组测序为RNA加工研究提供了额外信息，例如有关RNA结合蛋白的信息。在本章中，我们描述了一个详细的优化方案，用于构建高产高质量的降解组文库，以及NGS和数据挖掘程序。我们希望降解组方法将应用于非模式生物的进一步研究。
