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LC/MS/MS-Based Quantitative Assay for the Secondary Messenger Molecule, c-di-GMP

  • Yasuhiko Irie
  • Matthew R. ParsekEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1149)

Abstract

The secondary messenger molecule, 3′,5′-cyclic diguanosine monophosphate (c-di-GMP), controls various cellular processes in bacteria. Direct measurement of intracellular concentration of c-di-GMP is fast becoming an important tool for studying prokaryotic biology. Here, we describe a comprehensive extraction protocol from live bacteria and quantitative analysis using LC/MS/MS.

Key words

c-di-GMP LC/MS/MS Quantitative analysis Pseudomonas aeruginosa Nucleotide extraction 

Notes

Acknowledgments

The authors would like to thank Dale Whittington, Ross Lawrence, and the late Thomas Kalhorn for their expertise in LC/MS/MS. This work was supported in part by NIH grant MCB0822405 and AI061396-06 to MRP. YI was a UW CFF RDP Fellow.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Centre for Biomolecular Sciences, School of Molecular Medical SciencesUniversity of NottinghamNottinghamUK
  2. 2.Department of MicrobiologyUniversity of WashingtonSeattleUSA

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