NASC-seq monitors RNA synthesis in single cells
Lisa A. Jung,
Anton J.M. Larsson,
Oscar Andersson Forsman,
Michael Maximilian Lidschreiber,
Posted 17 Dec 2018
bioRxiv DOI: 10.1101/498667 (published DOI: 10.1038/s41467-019-11028-9)
Posted 17 Dec 2018
Sequencing of newly synthesized RNA can monitor transcriptional dynamics with great sensitivity and high temporal resolution, but is currently restricted to populations of cells. Here, we developed newly synthesized alkylation-dependent single-cell RNA sequencing (NASC-seq), to monitor both newly synthesized and pre-existing RNA in single cells. We validated the method on pre-alkylated exogenous spike-in RNA, and by demonstrating that more newly synthesized RNA was detected for genes with known high mRNA turnover. Importantly, NASC-seq reveals rapidly up- and down-regulated genes during the T-cell activation, and RNA sequenced for induced genes were essentially only newly synthesized. Moreover, the newly synthesized and pre-existing transcriptomes after T-cell activation were distinct confirming that we indeed could simultaneously measure gene expression corresponding to two time points in single cells. Altogether, NASC-seq is a powerful tool to investigate transcriptional dynamics and it will enable the precise monitoring of RNA synthesis at flexible time periods during homeostasis, perturbation responses and cellular differentiation.
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