Methods in Systems Biology

Contributors

Preface

Volume in series

Chapter one: Systems Biology Left and Right

1 A Fundamental Definition of Systems Biology

2 The Importance of the Integration of In Vivo and In Vitro Analyses

3 Alternative Definitions of Systems Biology

4 Different Types of Systems Biology

Chapter two: Mass Spectrometry in Systems Biology

1 Introduction

2 What is Mass Spectrometry?

3 Mass Spectrometer Configurations

4 The Benefits of Mass Spectrometry

5 Applications of Mass Spectrometry in Systems Biology

Chapter three: High-Throughput Quantification of Posttranslational Modifications In Situ by CA-FLIM

1 Introduction

2 Methods

3 Results and Discussion

4 Conclusion: Why Use CA-FLIM?

Appendix 1 Frequency Domain Fluorescence Lifetime Imaging Microscopy

Appendix 2 Global Analysis of FRET–FLIM Data

Appendix 3 Materials

Chapter four: Absorption Spectroscopy

1 Introduction

2 Theory of Absorption Spectroscopy

3 Hardware

4 Applications of UV–Visible Spectrometry

5 Perspective

Chapter five: A Strand-Specific Library Preparation Protocol for RNA Sequencing

1 Introduction

2 ssRNA-Seq Protocol

Chapter Six: Quantitative Real-Time PCR-Based Analysis of Gene Expression

1 Introduction

2 Primer Design

3 Primer Validation Experiments

4 Cell Culture-Based Isolation and Quantification of RNA

5 Reverse Transcription

Chapter seven: Quantification of Proteins and Their Modifications Using QconCAT Technology

1 Introduction

2 Expression and Purification of QconCAT Proteins

3 Preparation of Samples for Analysis

4 Mass Spectrometric Analysis

5 Quantification of Posttranslational Modifications

6 Replicate Analysis—Biological Versus Technical Replicates

7 Summary and General Considerations

Chapter eight: Mass Spectrometric-Based Quantitative Proteomics Using SILAC

1 Introduction

2 SILAC: The Method

3 General SILAC Protocol

4 Applications

5 Conclusion

Chapter nine: Nucleic Acid Programmable Protein Array

1 Introduction

2 Overview of NAPPA Chemistry

3 Array Production

4 Detection of Protein Expression

Chapter ten: Systems Biology of Recombinant Protein Production Using Bacillus megaterium

1 Introduction

2 Handling B. megaterium

3 Systems Biology of B. megaterium

Chapter eleven: Protein Production in Saccharomyces cerevisiae for Systems Biology Studies

1 Introduction

2 Comparison of Commonly Used Expression Systems

3 Comprehensive Libraries for Protein Production in S. cerevisiae

4 Protocols for Protein Expression and Purification from Tagged Collections of S. cerevisiae

5 Protein Analysis and Quantification

6 Protein Use in Proteomics and Enzyme Kinetics Measurements

7 Concluding Remarks

Chapter twelve: Towards a Full Quantitative Description of Yeast Metabolism

1 Introduction

2 Enzyme Kinetics for Systems Biology

3 Production and Purification of Isoenzymes

4 General Protocol for Enzymatic Assays

5 Assays for Measuring the Activities of the Glycolytic Isoenzymes of S. cerevisiae

6 Concluding Remarks

Chapter thirteen: Enzyme Kinetics for Systems Biology

1 Enzyme Kinetics for Systems Biology: Five Variations on the Theme

2 Three System Biology Approaches that Use Different Enzyme Kinetics

3 Biological Material

4 Protocols

5 Perspectives

Chapter fourteen: The Use of Continuous Culture in Systems Biology Investigations

1 Introduction

2 Experimental Considerations

3 Operation of the Permittistat

4 Sampling of Biomass for Proteome and Metabolome Analyses

Chapter fifteen: Sample Preparation Related to the Intracellular Metabolome of Yeast

1 Introduction

2 Sample Collection from Batch Cultures with Quenching of Intracellular Metabolism

3 Extraction of Polar and Nonpolar Metabolites from the Intracellular Metabolome

4 Targeted Quantification of Organic Acids Applying Gas Chromatography–Mass Spectrometry

Chapter sixteen: Plant Metabolomics and Its Potential for Systems Biology Research

1 An Introduction to Plant Metabolomics

2 Considerations in Plant Growth and Preparation of Plant Material

3 GC-TOF/MS Metabolite Profiling, Recommended Experimental Reference Protocols, and Data Preprocessing Approaches

4 HPLC–PDA–QTOFMS Metabolite Profiling, Recommended Experimental Reference Protocols, and Data Preprocessing Approaches

5 ¹H NMR: Experimental Reference Protocols and Data Preprocessing for Plant Metabolite Profiling

6 Concluding Remarks

Chapter seventeen: The Study of Mammalian Metabolism through NMR-based Metabolomics

1 Introduction

2 Tissue Extraction

3 Analysis of Biofluids by NMR Spectroscopy

4 A Brief Overview of Directly Measuring Metabolites in Mammalian Tissues by High-Resolution Magic Angle Spinning ¹H NMR Spectroscopy

5 Data Processing

6 Conclusions

Chapter eighteen: Building a Kinetic Model of Trehalose Biosynthesis in Saccharomyces cerevisiae

1 Introduction

2 Biological Background

3 Model Development

4 Results

5 Discussion

Chapter nineteen: Sustainable Model Building

1 Sustainable Model Building

2 How to Create Reusable Models

3 How to Implement Models in Standard Formats

4 How to Document and Annotate Models

5 Tools for Model Building and Annotation

6 How to Create and Draw a Simple Model

7 Annotations in Genome-Scale Network Reconstructions

Chapter twenty: From Reaction Networks to Information Flow—Using Modular Response Analysis to Track Information in Signaling Networks

1 Introduction

2 Modular Response Analysis

3 Conservation Analysis

4 From Reaction Schemes to Influence Networks Using a Monte Carlo Approach

5 Conclusion

Chapter twenty-one: Whole-Genome Metabolic Network Reconstruction and Constraint-Based Modeling

1 Introduction

2 Metabolic Network Reconstruction

3 Constraint-Based Modeling Methods

4 Summary

Chapter twenty-two: Hands-on Metabolism

1 Introduction

2 Elementary Flux Modes

3 Application

4 Conclusion

Appendix Reaction scheme of the photosynthate metabolism in metatool format (discussed in Section 3.2).

Chapter twenty-three: How to Obtain True and Accurate Rate-Values

1 Introduction

2 Quantification of Broth Amount in the Cultivation Vessel

3 Forgotten Processes

4 Detecting Forgotten Processes and Systematic Errors

5 Propagation of Statistical Errors: Accuracy of Calculated Rates

6 Experimental Design to Obtain True and Accurate R-Values

7 Mathematics of Reconciliation

8 Conclusion

Chapter twenty-four: A Practical Guide to Genome-Scale Metabolic Models and Their Analysis

1 Introduction

2 Genome-Scale Metabolic Models: Their Place in the Spectrum of Modeling Options

3 The Art of Making Genome-Scale Metabolic Models

4 Applications of Genome-Scale Metabolic Models

Chapter twenty-five: Supply–Demand Analysis

1 Introduction

2 The Functional Organization of Metabolism

3 Quantitative Analysis of Supply–Demand Systems

4 Generalized Supply–Demand Analysis

5 Experimental Applications of Supply–Demand Analysis

Chapter twenty-six: Modular Kinetic Analysis

1 Introduction

2 Description of the Method

3 Applications

Chapter twenty-seven: Quantitative Analysis of Flux Regulation Through Hierarchical Regulation Analysis

1 Introduction

2 Theory of Regulation Analysis

3 Experimental Tools for Regulation Analysis

4 Strategies of Flux Regulation

5 Concluding Remarks

Chapter twenty-eight: Origins of Stochastic Intracellular Processes and Consequences for Cell-to-Cell Variability and Cellular Survival Strategies

1 Cell-to-Cell Heterogeneity and Measurement Techniques

2 Theoretical Insights and Experimental Evidence

3 Beneficial and Detrimental Effects of Molecular Noise

4 Conclusion

Chapter twenty-nine: The SEEK

1 Introduction

2 The SEEK Platform

3 The Challenges of Data Management

4 The JERM Infrastructure

5 The SEEK Functionalities: Annotating and Linking Assets

6 Incentives for Sharing Data

7 The SEEK: Experiences

Chapter Thirty: Crossing the Boundaries

1 Introduction

2 Theoretical Management Strategies

3 Consideration of Real-World Examples

4 Conclusions

Author Index

Subject Index