Butt-seq: a new method for facile profiling of transcription
- Department of Biology, Howard Hughes Medical Institute, National Center for Behavioral Genomics, Brandeis University, Waltham, Massachusetts 02453, USA
- Corresponding author: rosbash{at}brandeis.edu
Abstract
A wide range of sequencing methods has been developed to assess nascent RNA transcription and resolve the single-nucleotide position of RNA polymerase genome-wide. These techniques are often burdened with high input material requirements and lengthy protocols. We leveraged the template-switching properties of thermostable group II intron reverse transcriptase (TGIRT) and developed Butt-seq (bulk analysis of nascent transcript termini sequencing), which can produce libraries from purified nascent RNA in 6 h and from as few as 10,000 cells—an improvement of at least 10-fold over existing techniques. Butt-seq shows that inhibition of the superelongation complex (SEC) causes promoter-proximal pausing to move upstream in a fashion correlated with subnucleosomal fragments. To address transcriptional regulation in a tissue, Butt-seq was used to measure the circadian regulation of transcription from fly heads. All the results indicate that Butt-seq is a simple and powerful technique to analyze transcription at a high level of resolution.
Keywords
- circadian rhythms
- nascent RNA
- RNA polymerase II pausing
- superelongation complex
- transcriptional profiling
- transcriptional regulation
Footnotes
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Supplemental material is available for this article.
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Article published online ahead of print. Article and publication date are online at http://www.genesdev.org/cgi/doi/10.1101/gad.350434.123.
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Freely available online through the Genes & Development Open Access option.
- Received January 14, 2023.
- Accepted April 20, 2023.
This article, published in Genes & Development, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.