Differential gene expression profiling in response to paraquat using a Drosophila Parkinson's disease model

Transcriptomic profiling using Drosophila heads reveals early gene expression responses to transient paraquat exposure. We performed RNAseq profiling of heads from the wild type (WT) Drosophila strain, Canton S (males only), that were fed either 2.5% sucrose (control) or 5 mM PQ in 2.5% sucrose for 12 h. Transcriptomic profiling was performed using independent biological replicates per feeding. Age-matched adult Canton S male flies were fed either 2.5% sucrose (control) or 5 mM PQ in 2.5% sucrose for 12 h and the heads were collected for RNA extraction. For paired-end library preparation, NEBNext mRNA Library Prep Reagent Set for Illumina was used and the library concentration was analyzed by utilizing a DNA 1000 Chip on an Agilent 2100 Bioanalyzer. Paired-end sequencing (25 million, 50-bp, paired-end reads) was performed using a 200 Cycle TruSeq SBS HS v4 Kit on an Illumina HiSeq2500 sequencer (Illumina, Inc., San Diego, CA, USA) at the HudsonAlpha Institute for Biotechnology, Huntsville, AL. TopHat v2.0 were used to map raw reads to the reference Drosophila melanogaster genome dm3

以果蝇头部为样本开展转录组谱分析，旨在揭示果蝇对百草枯（paraquat, PQ）短暂暴露的早期基因表达应答。本研究针对野生型（wild type, WT）果蝇坎顿S（Canton S）品系的雄性成虫头部进行RNA测序（RNA-seq）分析，实验分为两组：分别饲喂2.5%蔗糖溶液（对照组），或饲喂添加了5 mM PQ的2.5%蔗糖溶液，处理时长均为12小时，且每个饲喂组均设置独立生物学重复。本研究同步选取同龄的坎顿S品系雄性成虫，分别以2.5%蔗糖溶液（对照组）和含5 mM PQ的2.5%蔗糖溶液饲喂12小时后，收集其头部组织用于RNA提取。双端测序文库的制备采用适配Illumina平台的NEBNext mRNA文库制备试剂套装（NEBNext mRNA Library Prep Reagent Set for Illumina），并通过安捷伦2100生物分析仪（Agilent 2100 Bioanalyzer）搭配DNA 1000芯片（DNA 1000 Chip）检测文库浓度。测序工作在美国阿拉巴马州亨茨维尔市哈德森阿尔法生物技术研究所（HudsonAlpha Institute for Biotechnology, Huntsville, AL, USA）的Illumina HiSeq2500测序仪（Illumina, Inc., San Diego, CA, USA）上完成，采用200循环TruSeq SBS HS v4测序试剂盒进行双端测序，产出2500万条50 bp双端读段。最后使用TopHat v2.0软件将原始读段比对至黑腹果蝇（Drosophila melanogaster）参考基因组dm3。