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NASC-seq monitors RNA synthesis in single cells

By Gert-Jan Hendriks, Lisa A. Jung, Anton J.M. Larsson, Oscar Andersson Forsman, Michael Maximilian Lidschreiber, Katja Lidschreiber, Patrick Cramer, Rickard Sandberg

Posted 17 Dec 2018
bioRxiv DOI: 10.1101/498667 (published DOI: 10.1038/s41467-019-11028-9)

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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