RNA测序（英语：RNA Sequencing，简称RNA-Seq，也被称为全转录物组鸟枪法测序 Whole Transcriptome Shotgun Sequencing，简称WTSS）是基于第二代测序技术的转录组学研究方法。RNA测序是使用第二代测序的能力，在给定时刻从一个基因组中，揭示RNA的存在和数量的一个快照的技术。

首先提取生物样品的全部转录的RNA，然后反转录为c-DNA后进行的二代高通量测序，在此基础上进行片段的重叠组装，从而可得到一个个的转录本。进而可以形成对该生物样品当前发育状态的基因表达状况的全局了解（global）。进一步说，若和下一阶段的生物样品的RNA-Seq转录组进行比较，则可以得到全部的（在转录层面）基因表达的上调及下调--这就形成了表达谱，针对关键基因则可以形成你要想要的pathway的构建。

RNA-seq (RNA sequencing), also called whole transcriptome shotgun sequencing (WTSS), uses next-generation sequencing (NGS) to reveal the presence and quantity of RNA in a biological sample at a given moment in time.

RNA-Seq is used to analyze the continually changing cellular transcriptome. Specifically, RNA-Seq facilitates the ability to look at alternative gene spliced transcripts, post-transcriptional modifications, gene fusion, mutations/SNPs and changes in gene expression. In addition to mRNA transcripts, RNA-Seq can look at different populations of RNA to include total RNA, small RNA, such as miRNA, tRNA, and ribosomal profiling. RNA-Seq can also be used to determine exon/intron boundaries and verify or amend previously annotated 5’ and 3’ gene boundaries.