RNA Stable Isotope Probing (RNA-SIP)

Noor Ul Huda Ghori, Benjamin Moreira-Grez, Paton Vuong, Ian Waite, Tim Morald, Michael Wise, Andrew S. Whiteley

Research output: Chapter in Book/Conference paperChapter

Abstract

Stable isotope probing is a combined molecular and isotopic technique used to probe the identity and function of uncultivated microorganisms within environmental samples. Employing stable isotopes of common elements such as carbon and nitrogen, RNA-SIP exploits an increase in the buoyant density of RNA caused by the active metabolism and incorporation of heavier mass isotopes into the RNA after cellular utilization of labeled substrates pulsed into the community. Labeled RNAs are subsequently separated from unlabeled RNAs by density gradient centrifugation followed by identification of the RNAs by sequencing. Therefore, RNA stable isotope probing is a culture-independent technique that provides simultaneous information about microbiome community, composition and function. This chapter presents the detailed protocol for performing an RNA-SIP experiment, including the formation, ultracentrifugation, and fractional analyses of stable isotope-labeled RNAs extracted from environmental samples.

Original languageEnglish
Title of host publicationStable Isotope Probing
Subtitle of host publicationMethods and Protocols
EditorsMarc G. Dumont, Marcela Hernández García
Place of PublicationUSA
PublisherHumana Press
Pages31-44
Number of pages14
ISBN (Electronic)9781493997213
ISBN (Print)9781493997206
DOIs
Publication statusPublished - 1 Jan 2019

Publication series

NameMethods in Molecular Biology
Volume2046
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

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Ghori, N. U. H., Moreira-Grez, B., Vuong, P., Waite, I., Morald, T., Wise, M., & Whiteley, A. S. (2019). RNA Stable Isotope Probing (RNA-SIP). In M. G. Dumont, & M. Hernández García (Eds.), Stable Isotope Probing : Methods and Protocols (pp. 31-44). (Methods in Molecular Biology; Vol. 2046). USA: Humana Press. https://doi.org/10.1007/978-1-4939-9721-3_3