The emergence of next generation sequencing (NGS) has facilitated the high-throughput sequencing of entire cellular repertoires of RNA transcripts. Subsequently, the sequencing of RNA molecules (RNA-Seq) is rapidly becoming the method of choice for global gene expression profiling as it can provide a very precise measurement of levels of RNA transcripts. RNA sequencing can also be applied to investigating alternatively spliced transcripts, post-transcriptional modifications, gene fusion events and transcribed mutations/SNPs. Numerous RNA-Seq studies have already facilitated a massive shift in our view of the extent and complexity of mammalian gene regulation. In this presentation, I will describe RNA-Seq, how it can be applied to answer novel questions in reproductive biology, the challenges associated with RNA-Seq data analysis, and recent advances facilitated by innovative applications of RNA-Seq. Throughout this talk I will draw upon examples from our recent profiling of the human placental transcriptome to demonstrate the power and pitfalls of RNA-Seq.