Külahoglu Canan, Bräutigam Andrea
iGrad Plant, Heinrich Heine University Düsseldorf, Universitätsstrasse 1, 40225, Düsseldorf, Germany.
Methods Mol Biol. 2014;1158:71-91. doi: 10.1007/978-1-4939-0700-7_5.
RNA-seq has emerged as the technology of choice to quantify gene expression. This technology is a convenient accurate tool to quantify diurnal changes in gene expression, gene discovery, differential use of promoters, and splice variants for all genes expressed in a single tissue. Thus, RNA-seq experiments provide sequence information and absolute expression values about transcripts in addition to relative quantification available with microarrays or qRT-PCR. The depth of information by sequencing requires careful assessment of RNA intactness and DNA contamination. Although the RNA-seq is comparatively recent, a standard analysis framework has emerged with the packages of Bowtie2, TopHat, and Cufflinks. With rising popularity of RNA-seq tools have become manageable for researchers without much bioinformatical knowledge or programming skills. Here, we present a workflow for a RNA-seq experiment from experimental planning to biological data extraction.
RNA测序已成为定量基因表达的首选技术。这项技术是一种方便且准确的工具，可用于定量基因表达的昼夜变化、基因发现、启动子的差异使用以及单个组织中所有表达基因的剪接变体。因此，RNA测序实验除了提供微阵列或qRT-PCR可获得的相对定量信息外，还能提供有关转录本的序列信息和绝对表达值。测序所提供的信息深度要求对RNA完整性和DNA污染进行仔细评估。尽管RNA测序相对较新，但已经出现了一个由Bowtie2、TopHat和Cufflinks软件包组成的标准分析框架。随着RNA测序工具越来越受欢迎，没有太多生物信息学知识或编程技能的研究人员也能够操作这些工具。在这里，我们展示了一个从实验规划到生物数据提取的RNA测序实验工作流程。
