RT Journal Article
SR Electronic
T1 Single cell profiling of total RNA using Smart-seq-total
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.06.02.131060
DO 10.1101/2020.06.02.131060
A1 Alina Isakova
A1 Norma Neff
A1 Stephen R. Quake
YR 2020
UL http://biorxiv.org/content/early/2020/06/03/2020.06.02.131060.abstract
AB The ability to interrogate total RNA content of single cells would enable better mapping of the transcriptional logic behind emerging cell types and states. However, current RNA-seq methods are unable to simultaneously monitor both short and long, poly(A)+ and poly(A)-transcripts at the single-cell level, and thus deliver only a partial snapshot of the cellular RNAome. Here, we describe Smart-seq-total, a method capable of assaying a broad spectrum of coding and non-coding RNA from a single cell. Built upon the template-switch mechanism, Smart-seq-total bears the key feature of its predecessor, Smart-seq2, namely, the ability to capture full-length transcripts with high yield and quality. It also outperforms current poly(A)–independent total RNA-seq protocols by capturing transcripts of a broad size range, thus, allowing us to simultaneously analyze protein-coding, long non-coding, microRNA and other non-coding RNA transcripts from single cells. We used Smart-seq-total to analyze the total RNAome of human primary fibroblasts, HEK293T and MCF7 cells as well as that of induced murine embryonic stem cells differentiated into embryoid bodies. We show that simultaneous measurement of non-coding RNA and mRNA from the same cell enables elucidation of new roles of non-coding RNA throughout essential processes such as cell cycle or lineage commitment. Moreover, we show that cell types can be distinguished based on the abundance of non-coding transcripts alone.Competing Interest StatementThe authors have declared no competing interest.