In situ 10-cell RNA sequencing in tissue and tumor biopsy samples [article]

Shambhavi Singh, Lixin Wang, Dylan L. Schaff, Matthew D. Sutcliffe, Alex F. Koeppel, Jungeun Kim, Suna Onengut-Gumuscu, Kwon-Sik Park, Hui Zong, Kevin A. Janes
2018 bioRxiv   pre-print
Single-cell transcriptomic methods classify new and existing cell types very effectively, but alternative approaches are needed to quantify the individual regulatory states of cells in their native tissue context. We combined the tissue preservation and single-cell resolution of laser capture with an improved preamplification procedure enabling RNA sequencing of 10 microdissected cells. This in situ 10-cell RNA sequencing (10cRNA-seq) can exploit fluorescent reporters of cell type in
more » ... engineered mice and is compatible with freshly cryoembedded clinical biopsies from patients. Through recombinant RNA spike-ins, we estimate dropout-free technical reliability as low as ~250 copies and a 50% detection sensitivity of ~45 copies per 10-cell reaction. By using small pools of microdissected cells, 10cRNA-seq improves per-cell reliability and sensitivity beyond existing approaches for single-cell RNA sequencing (scRNA-seq). Accordingly, in multiple tissue and tumor settings, we observe 1.5-2-fold increases in genes detected and overall alignment rates compared to scRNA-seq. Combined with existing approaches to deconvolve small pools of cells, 10cRNA-seq offers a reliable, unbiased, and sensitive way to measure cell-state heterogeneity in tissues and tumors.
doi:10.1101/444182 fatcat:ydkxypcwhnddve7zws3ue5nnve