Determining mRNA decay rates using RNA approach to equilibrium sequencing (RATE-seq)

Farah Abdul-Rahman, David Gresham

Research output: Chapter in Book/Report/Conference proceedingChapter


RATE-seq is a 4-thiouracil (4-tU)-based method that enables the in vivo measurement of transcriptome-wide RNA degradation rates. 4-tU is an analog of uracil that is rapidly incorporated into newly synthesized RNA and facilitates the conjugation of a biotinylated molecule containing a reactive thiol group. The biotinylated RNA can then be fractionated from the unlabeled RNA with streptavidin magnetic beads. By adding 4-tU to a culture of cells growing in steady-state conditions, fractionating the labeled population of RNA at multiple time points following 4-tU addition, and quantifying the abundance of newly transcribed RNAs using RNAseq, it is possible to estimate the degradation rates of all transcripts in a single experiment. The analysis of the RATE-seq data entails normalization of RNAseq libraries to thiolated RNA spike-ins and nonlinear model fitting to estimate the degradation rate constant for each RNA species.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Number of pages10
ISBN (Print)9781493975402
StatePublished - 2018

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745


  • 4-thiouracil
  • Metabolic labeling
  • RNA degradation
  • RNA stability
  • RNA turnover

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics


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