RNA sequencing: Platform selection, experimental design, and data interpretation

Detailed characterization of cellular RNA facilitates the design of nucleic acid therapeutics and interpretation of experimental data. Since the first reports of next generation sequencing (NGS) technology-based RNA sequencing (RNA-seq) (Nagalakshmi et al., 2008; Wilhelm, et al., 2008), rapid advances in experimental protocol development and data acquisition and analysis have brought comprehensive sequencing of cellular RNA within reach of most laboratories (Martin and Wang, 2011; Ozsolak and Milos, 2011). While RNA-seq is becoming widely used for laboratory research and clinical studies, the field is still new and strategies for successfully applying RNA-seq to a given experimental problem are often obscure. The goal of this perspective is to introduce some of the choices confronted during RNA sequencing and analysis.