Abstract
Small regulatory RNAs (sRNAs) are ubiquitous regulatory molecules expressed in living cells. In prokaryotes, sRNAs usually bind to target mRNAs to either promote their degradation or interfere with translation initiation. Because a single sRNA can regulate a considerable number of target mRNAs, we seek to identify those targets rapidly and reliably. Here, we present a robust method based on the co-purification of target mRNAs bound to MS2-tagged sRNAs expressed in vivo. After purification of the tagged-sRNA, we use RNAseq to determine the identity of all RNA interacting partners and their enrichment level. We describe how to analyze the RNAseq data through the Galaxy Project Platform bioinformatics tools to identify new mRNA targets. This technique is applicable to most sRNAs of E. coli and Salmonella.
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Acknowledgments
This work was funded by an operating grant from the Canadian Institutes of Health Research (CIHR) to EM. M.-C.C. holds an Alexander Graham Bell Doctoral scholarship from the Natural Sciences and Engineering Research Council of Canada (NSERC).
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Carrier, MC., Laliberté, G., Massé, E. (2018). Identification of New Bacterial Small RNA Targets Using MS2 Affinity Purification Coupled to RNA Sequencing. In: Arluison, V., Valverde, C. (eds) Bacterial Regulatory RNA. Methods in Molecular Biology, vol 1737. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7634-8_5
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DOI: https://doi.org/10.1007/978-1-4939-7634-8_5
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