RNA structure probing to characterize RNA-protein interactions on a low abundance pre-mRNA in living cells

In vivo RNA structure analysis has become a powerful tool in molecular biology, largely due to the coupling of an increasingly diverse set of chemical approaches with high-throughput sequencing. This has resulted in a transition from single target to transcriptome-wide approaches. However, these met...

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Bibliographic Details
Published in:RNA (Cambridge) 2021-03, Vol.27 (3), p.343-358
Main Authors: Bubenik, Jodi L, Hale, Melissa, McConnell, Ona, Wang, Eric, Swanson, Maurice S, Spitale, Robert, Berglund, J Andrew
Format: Article
Language:English
Subjects:
Gene expression
Method
Next-generation sequencing
Protein interaction
Reverse transcription
Splicing factors
Transcriptomes
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Summary:In vivo RNA structure analysis has become a powerful tool in molecular biology, largely due to the coupling of an increasingly diverse set of chemical approaches with high-throughput sequencing. This has resulted in a transition from single target to transcriptome-wide approaches. However, these methods require sequencing depths that preclude studying low abundance targets, which are not sufficiently captured in transcriptome-wide approaches. Here we present a ligation-free method to enrich for low abundance RNA sequences, which improves the diversity of molecules analyzed and results in improved analysis. In addition, this method is compatible with any choice of chemical adduct or read-out approach. We utilized this approach to study an autoregulated event in the pre-mRNA of the splicing factor, muscleblind-like splicing regulator 1 (MBNL1).
ISSN:1355-8382
1469-9001
DOI:10.1261/rna.077263.120