scNMT-seq of pancreatic organoids - RNA

Single-cell nucleosome, methylome and transcriptome (scNMT) sequencing is a recently developed method that allows multiomics profiling of single cells. In this scNMT protocol, we describe profiling of cells from mouse brain and pancreatic organoids, using liquid handling platforms to increase throughput from 96-well to 384-well plate format. Our approach miniaturizes reaction volumes and incorporates the latest Smart-seq3 protocol to obtain a higher number of detected genes and genomic DNA (gDNA) CpG sites per cell. We outline normalization steps to optimally distribute per-cell sequencing depth. Primary pancreatic acinar cells were isolated by digestive dissociation of pancreatic tissue and subsequent density gradient centrifugation. After cultivation as 3D matrigel embedded organoids for 9 days, the cells were harvested and subjected to the miniaturized scNMT-seq protocol.