Abstract
Second-generation RNA-based fluorescent biosensors have been developed that enable flow cytometry experiments to monitor the population dynamics of c-di-GMP signaling in live bacteria. These experiments are high-throughput, provide information at the single-cell level, and can be performed on cells grown in complex media and/or under anaerobic conditions. Here, we describe flow cytometry methods for three applications: (1) high-throughput screening for diguanylate cyclase activity, (2) analyzing c-di-GMP levels under anaerobic conditions, and (3) monitoring cell population dynamics of c-di-GMP levels upon environmental changes. These methods showcase RNA-based fluorescent biosensors as versatile tools for studying c-di-GMP signaling in bacteria.
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Acknowledgment
The work on which this chapter is based was supported by NIH grant DP2 OD008677 (to M.C.H.). The authors thank Zachary Hallberg for assistance with figures.
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Yeo, J., Wang, X.C., Hammond, M.C. (2017). Live Flow Cytometry Analysis of c-di-GMP Levels in Single Cell Populations. In: Sauer, K. (eds) c-di-GMP Signaling. Methods in Molecular Biology, vol 1657. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7240-1_10
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DOI: https://doi.org/10.1007/978-1-4939-7240-1_10
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