Next-generation sequencing facilitates quantitative analysis of wild and acute renal failure mice

The goals of this study are to compare NGS-derived renal transcriptome profiling (RNA-seq) to microarray and to evaluate protocols for optimal high-throughput data analysis. Methods: Renal mRNA profiles of 21-day-old Wild and Acute renal failure mice were generated by deep sequencing, in triplicate. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays.

本研究旨在对比基于下一代测序（Next-Generation Sequencing, NGS）的肾脏转录组谱分析（RNA-seq）与微阵列技术，并优化高通量数据分析的最优方案。方法：选取21日龄野生型小鼠与急性肾衰竭小鼠，通过三重生物学重复的深度测序获取其肾脏mRNA表达谱。对通过质量过滤的测序读段（reads），在转录本异构体水平采用两种分析流程：先使用Burrows–Wheeler Aligner（BWA）后结合方差分析（ANOVA），以及先使用TopHat后进行Cufflinks分析。采用TaqMan探针法与SYBR Green染料法开展qRT-PCR验证实验。