Single-Cell mRNA Sequencing of the Mouse Brain Vasculature

In this chapter, we describe a method for analyzing the vasculature of the mouse brain using single-cell transcriptomics. More specifically, we focus on the use of fluorescence-activated cell sorting (FACS) for selection of the cells of interest and sorting of these cells in a 384-well format, allowing for enrichment for the cells being studied. Furthermore, we outline the Smart-Seq2 single-cell library construction method for transforming single-cell mRNA into Illumina sequencing compatible libraries. As single-cell sequencing is still a costly technology, we take special care to describe strategies to include many quality control steps. Finally, we touch upon techniques to convert the sequencing data into a meaningful biological readout. The methods reported in this chapter can be expanded toward other tissues and will prove useful also for the study of different cell types beyond adult brain vasculature.