An ongoing technological revolution is continually improving our ability to carry out very high-resolution studies of gene expression patterns. Current technology enables the global gene expression profiles of single cells to be defined, facilitating dissection of heterogeneity in cell populations that was previously hidden. In contrast to gene expression studies that use bulk RNA samples and provide only a virtual average of the diverse constituent cells, single-cell studies enable the molecular distinction of all cell types within a complex population mix, such as a tumour or developing organ. For instance, single-cell gene expression profiling has contributed to improved understanding of how histologically identical, adjacent cells make different differentiation decisions during development. Beyond development, single-cell gene expression studies have enabled the characteristics of previously known cell types to be more fully defined and facilitated the identification of novel categories of cells, contributing to improvements in our understanding of both normal and disease-related physiological processes and leading to the identification of new treatment approaches. Although limitations remain to be overcome, technology for the analysis of single-cell gene expression patterns is improving rapidly and beginning to provide a detailed atlas of the gene expression patterns of all cell types in the human body.
A microfluidics system is used to make microdroplets, which contain cells mixed with beads that are encapsulated in oil. Each bead has oligonucleotides that are uniquely barcoded for that bead and are in a solution that contains a mild detergent, which lyses the cells after mixing. The RNAs from the lysed cell anneal to the bead oligonucleotides, and subsequent reverse transcription incorporates the bead-specific barcode into the cDNA, thereby allowing the sequences of those cDNAs to be assigned to a specific cell. scRNA-seq, single-cell RNA sequencing.
- RNA sequencing of single cells (scRNA-seq) enables the global gene expression patterns of individual cells to be defined.
- Almost all tissues and organs include a heterogeneous mix of cell types; the heterogeneity of these cell populations can be defined through the use of scRNA-seq.
- scRNA-seq can fully define the expression of transcription factors, growth factors, receptors, solute transporters and other proteins for each cell type present, providing insights into cell function and cell–cell crosstalk.
- scRNA-seq is an increasingly powerful tool for the analysis of development as well as normal and disease processes.