Time course analysis of gene expression during hypoxia in several S. cerevisiae mutant strains using RNA-Seq. Time course analysis of gene expression during hypoxia in several S. cerevisiae mutant strains using RNA-Seq

We used RNA-seq to monitor mRNA levels of all genes in response to hypoxia in several mutant strains of the yeast, S. cerevisiae (S288C background with wildtype HAP1). To gain insights into how gene expression changes over time, cells were subjected to 100% nitrogen gas and collected after 0,5,10,30,60,120,180, and 240 minutes. Total RNA was extracted and mRNAs were enriched by polyA selection. The cDNA was prepared into a sequencing library, multiplexed and single-end sequenced by an Illumina HiSeq 2500 sequencer. After mapping with Tophat2, the number of reads per feature was calculated using HTSeq. Overall design: RNA-seq analysis of eight time points of a yeast strain grown in hypoxia. The time course course is repeated in different mutant strains.

本研究采用RNA测序（RNA-seq）技术，监测酿酒酵母（Saccharomyces cerevisiae，简称S. cerevisiae，S288C背景且携带野生型HAP1）多个突变菌株中所有基因的mRNA水平对低氧的响应情况。为探究基因表达随时间的动态变化规律，将酵母细胞置于100%氮气环境中培养，并分别于0、5、10、30、60、120、180及240分钟时收集细胞样本。总RNA经提取后，通过polyA选择（polyA selection）法富集mRNA。将反转录得到的cDNA构建为测序文库，经多重复合标记后使用Illumina HiSeq 2500测序仪完成单端测序。利用Tophat2完成序列比对后，采用HTSeq软件计算每个基因特征的读段计数。整体实验设计：对低氧环境下培养的酵母菌株的8个时间点样本开展RNA-seq分析，该时间进程实验在不同的突变菌株中重复开展。