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Isotopically Nonstationary 13C Metabolic Flux Analysis

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Systems Metabolic Engineering

Part of the book series: Methods in Molecular Biology ((MIMB,volume 985))

Abstract

13C metabolic flux analysis (MFA) is a powerful approach for quantifying cell physiology based upon a combination of extracellular flux measurements and intracellular isotope labeling measurements. In this chapter, we present the method of isotopically nonstationary 13C MFA (INST-MFA), which is applicable to systems that are at metabolic steady state, but are sampled during the transient period prior to achieving isotopic steady state following the introduction of a 13C tracer. We describe protocols for performing the necessary isotope labeling experiments, for quenching and extraction of intracellular metabolites, for mass spectrometry (MS) analysis of metabolite labeling, and for computational flux estimation using INST-MFA. By combining several recently developed experimental and computational techniques, INST-MFA provides an important new platform for mapping carbon fluxes that is especially applicable to animal cell cultures, autotrophic organisms, industrial bioprocesses, high-throughput experiments, and other systems that are not amenable to steady-state 13C MFA experiments.

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Acknowledgements

This work was supported by NSF EF-1219603. LJJ was supported by a GAANN fellowship from the US Department of Education under grant number P200A090323.

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Authors and Affiliations

  1. Department of Chemical and Biomolecular Engineering, Vanderbilt University, Nashville, TN, USA

    Lara J. Jazmin & Jamey D. Young

Authors
  1. Lara J. Jazmin
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  2. Jamey D. Young
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Correspondence to Jamey D. Young .

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Editors and Affiliations

  1. Cockrell School of Engineering, Dept. of Chemical Engineering, The University of Texas at Austin, E. Dean Keeton Street 200, Austin, 78712, Texas, USA

    Hal S. Alper

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Jazmin, L.J., Young, J.D. (2013). Isotopically Nonstationary 13C Metabolic Flux Analysis. In: Alper, H. (eds) Systems Metabolic Engineering. Methods in Molecular Biology, vol 985. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-299-5_18

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  • DOI: https://doi.org/10.1007/978-1-62703-299-5_18

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-298-8

  • Online ISBN: 978-1-62703-299-5

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