Single-cell RNA-sequencing of Platynereis dumerilii larval brain cells

Understanding cell type identity in complex tissues or organisms requires integration of each cell’s expression profile with its spatial location within the tissue under study. We developed a high-throughput method that combines in vitro single-cell RNA-sequencing with a gene expression atlas to map single cells back to their location within the tissue of interest. We used the developing brain of a marine annelid, Platynereis dumerilii that is an important model system for studying bilaterian brain evolution, to benchmark our approach. To generate the single-cell mRNA-sequencing data, P. dumerilii larval brains were dissociated, followed by cell capture, cDNA synthesis and amplification on the C1 Single-Cell Auto Prep IFC for 10-17 um cells (Fluidigm). Sequencing libraries were produced using Nexera XT DNA kit (Illumina). In total, we sequenced 141 samples, of which 91 correspond to single, alive cells (as judged by visual inspection of the captured cells) with the remainder consisting of a variety of single dead cells (n=3), wells containing extracellular matrix contaminants (n=4) or multiple cells (n=11), as well as a negative controls where no cells were observed (n=32). For this dataset, we achieved 90% success rate for the spatial mapping of the single-cell RNA-seq data to P. dumerilii brain atlas.