ABSTRACTThe past four decades have yielded advances in molecular biology allowing detailed characterization of the cellular genome and the transcriptome: the complete set of RNA species transcribed by a cell or tissue. Through transcriptomics and next‐generation sequencing, we can now attain an unprecedented level of detail in understanding cellular phenotypes through examining the genes expressed in specific physiological and pathological states. In this review, we provide an overview of transcriptomics and RNA‐sequencing in the analysis of whole tissue and single cells. We describe the techniques and pitfalls involved in the isolation and sequencing of single cells, and what additional benefits this application can provide. Finally, we look to how these technologies are being applied in pulmonary research, and how they may translate in the near future into clinical practice.