转录组
生物
RNA序列
核糖核酸
计算生物学
基因
寄主(生物学)
核糖体RNA
基因表达
遗传学
作者
Alexander J. Westermann,Jörg Vogel
出处
期刊:Methods in molecular biology
日期:2018-01-01
卷期号:: 59-75
被引量:51
标识
DOI:10.1007/978-1-4939-7634-8_4
摘要
Transcriptomics, i.e., the quantification of cellular RNA transcripts, is a powerful way to gauge the physiological state of either bacterial or eukaryotic cells under a given condition. However, traditional approaches were unsuitable to measure the abundance of transcripts across kingdoms, which is relevant for biological processes such as bacterial infections of mammalian host cells. This changed with the establishment of “Dual RNA-seq,” which profiles gene expression simultaneously in an infecting bacterium and its infected host. Here, we describe a detailed Dual RNA-seq protocol optimized for—but not restricted to—the study of human cell culture models infected with the Gram-negative model pathogen Salmonella Typhimurium. Furthermore, we provide experimental data demonstrating the benefits of some of the key steps of this protocol, including transcriptome stabilization (RNA fixation), FACS-based enrichment of invaded cells, and double rRNA depletion. While our focus is on data generation, we also include a section describing suitable computational methods to analyze the obtained datasets.
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