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Tracking Distinct RNA Populations Using Efficient and Reversible Covalent Chemistry.

ABSTRACT: We describe a chemical method to label and purify 4-thiouridine (s(4)U)-containing RNA. We demonstrate that methanethiosulfonate (MTS) reagents form disulfide bonds with s(4)U more efficiently than the commonly used HPDP-biotin, leading to higher yields and less biased enrichment. This increase in efficiency allowed us to use s(4)U labeling to study global microRNA (miRNA) turnover in proliferating cultured human cells without perturbing global miRNA levels or the miRNA processing machinery. This improved chemistry will enhance methods that depend on tracking different populations of RNA, such as 4-thiouridine tagging to study tissue-specific transcription and dynamic transcriptome analysis (DTA) to study RNA turnover.

SUBMITTER: Duffy EE

PROVIDER: S-EPMC4560836 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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Tracking Distinct RNA Populations Using Efficient and Reversible Covalent Chemistry.

Duffy Erin E EE Rutenberg-Schoenberg Michael M Stark Catherine D CD Kitchen Robert R RR Gerstein Mark B MB Simon Matthew D MD

Molecular cell 20150901 5

We describe a chemical method to label and purify 4-thiouridine (s(4)U)-containing RNA. We demonstrate that methanethiosulfonate (MTS) reagents form disulfide bonds with s(4)U more efficiently than the commonly used HPDP-biotin, leading to higher yields and less biased enrichment. This increase in efficiency allowed us to use s(4)U labeling to study global microRNA (miRNA) turnover in proliferating cultured human cells without perturbing global miRNA levels or the miRNA processing machinery. Thi ...[more]

PMID: 26340425

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Tracking distinct RNA populations using efficient and reversible covalent chemistry

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