Metabolic Flux Analysis, 2014 Methods and Protocols Methods in Molecular Biology Series, Vol. 1191

Metabolic Flux Analysis: Methods and Protocols opens up the field of metabolic flux analysis to those who want to start a new flux analysis project but are overwhelmed by the complexity of the approach. Metabolic flux analysis emerged from the current limitation for the prediction of metabolic fluxes from a measured inventory of the cell. Divided into convenient thematic parts, topics in this essential volume include the fundamental characteristics of the underlying networks, the application of quantitative metabolite data and thermodynamic principles to constrain the solution space for flux balance analysis (FBA), the experimental toolbox to conduct different types of flux analysis experiments, the processing of data from ¹³C experiments and three chapters that summarize some recent key findings. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols and notes on troubleshooting and avoiding known pitfalls.

Authoritative and easily accessible, Metabolic Flux Analysis: Methods and Protocols presents protocols that cover a range of relevant organisms currently used in the field, providing a solid basis to anybody interested in the field of metabolic flux analysis.

Part I. Models - Stoichiometry, Network Generation

1. Stoichiometric Modelling of Microbial Metabolism

                Lars Kuepfer

2. Tapping the Wealth of Microbial Data in High-Throughput Metabolic Model Reconstruction

                Ric Colasanti, Janaka N. Edirisinghe, Tahmineh Khazaei, José P. Faria, Sam Seaver, Fangfang Xia, and Christopher Henry

Part II. Thermodynamics - A Step Beyond MFA

3. Constraining the Flux Space using Thermodynamics and Integration of Metabolomics Data

                Keng Cher Soh and Vassily Hatzimanikatis

4. NExT - Integration of Thermodynamic Constraints and Metabolomics Data into a Metabolic Network

                Verónica Sofía Martínez and Lars Keld Nielsen

Part III. Experiments - Design Considerations, Fermentation, Analytics

5. Customization of 13C-MFA Strategy According to Cell Culture System

                Lake-Ee Quek and Lars Keld Nielsen

6. Quantitative Metabolomics Using ID-MS

                Aljoscha S. Wahl, Reza Maleki Seifar, Angela ten Pierick, Cor Ras, Jan van Dam, Joseph J. Heijnen, and Walter M. Van Gulik

7. Determining the Biomass Composition of a Sponge Holobiont for Flux Analysis

                Jabin Watson, Bernard Degnan, Sandie Degnan, and Jens O. Krömer

8. Successful Down-Sizing for High-Throughput 13C-MFA Applications

                Birgitta E. Ebert and Lars M. Blank

9. Labelling Analysis for 13C MFA Using NMR Spectroscopy

                Paula Jouhten and Hannu Maaheimo

10. GC-MS Based 13C Metabolic Flux Analysis

                Judith Becker and Christoph Wittman

11. Dynamic Cell Respiration & CO2 Labeling Analysis Using Membrane-Inlet Mass Spectrometry

                Tae Hoon Yang

Part IV. 13C Fluxomics Data Processing

12. Correction of MS Data for Naturally Occurring Isotopes in Isotope Labelling Experiments

                Pierre Millard, Fabien Letisse, Serguei Sokol, and Jean-Charles Portais

13. Steady-State 13C Fluxomics Using OpenFLUX

                Lake-Ee Quek and Lars Keld Nielsen

14. Flux Visualization Using VANTED/FluxMap

                Christian Krach, Astrid Junker, Hendrik Rohn, Falk Schreiber, and Björn H. Junker

Part V. Examples

15. Metabolic Flux Analysis for Escherichia coli by Flux Balance Analysis

                Yu Matsuoka and Kazuyuki Shimizu

16. 13C-Metabolic Flux Analysis for Escherichia coli

                Yu Matsuoka and Kazuyuki Shimizu

17. 3C-Based Metabolic Flux Analysis of Recombinant Pichia pastoris

                Pau Ferrer and Joan Albiol

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts