用于通过高通量测序分析小RNA的cDNA文库构建。

cDNA library generation for the analysis of small RNAs by high-throughput sequencing.

作者信息

Gebetsberger Jennifer, Fricker Roger, Polacek Norbert

机构信息

Department of Chemistry and Biochemistry, Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.

出版信息

Methods Mol Biol. 2015;1296:139-49. doi: 10.1007/978-1-4939-2547-6_13.

DOI:10.1007/978-1-4939-2547-6_13

PMID:25791597

Abstract

The RNome of a cell is highly diverse and consists besides messenger RNAs (mRNAs), transfer RNAs (tRNAs), and ribosomal RNAs (rRNAs) also of other small and long transcript entities without apparent coding potential. This class of molecules, commonly referred to as non-protein-coding RNAs (ncRNAs), is involved in regulating numerous biological processes and thought to contribute to cellular complexity. Therefore, much effort is put into their identification and further functional characterization. Here we provide a cost-effective and reliable method for cDNA library construction of small RNAs in the size range of 20-500 residues. The effectiveness of the described method is demonstrated by the analysis of ribosome-associated small RNAs in the eukaryotic model organism Trypanosoma brucei.

摘要

细胞的RNA组高度多样，除信使RNA（mRNA）、转运RNA（tRNA）和核糖体RNA（rRNA）外，还包括其他无明显编码潜能的小转录本和长转录本实体。这类分子通常被称为非蛋白质编码RNA（ncRNA），参与调节众多生物学过程，并被认为有助于细胞复杂性。因此，人们投入了大量精力对其进行鉴定和进一步的功能表征。在此，我们提供了一种经济高效且可靠的方法，用于构建长度在20 - 500个残基范围内的小RNA的cDNA文库。通过对真核模式生物布氏锥虫中与核糖体相关的小RNA进行分析，证明了所描述方法的有效性。