Synthetic Biology and Metabolic Engineering in Plants and Microbes Part B: Metabolism in Plants

1st Edition, Volume 576 - July 28, 2016
Editor: Sarah E O'Connor
Language: English
Hardback ISBN:
9 7 8 - 0 - 1 2 - 8 0 4 5 3 9 - 8
eBook ISBN:
9 7 8 - 0 - 1 2 - 8 1 1 1 2 3 - 9

Synthetic Biology and Metabolic Engineering in Plants and Microbes, Part B, the latest volume in the Methods in Enzymology series, continues the legacy of this premier serial wit… Read more

Synthetic Biology and Metabolic Engineering in Plants and Microbes Part B: Metabolism in Plants

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Synthetic Biology and Metabolic Engineering in Plants and Microbes, Part B, the latest volume in the Methods in Enzymology series, continues the legacy of this premier serial with quality chapters authored by leaders in the field.

This volume covers research methods, synthetic biology, and metabolic engineering in plants and microbes, and includes sections on such topics as the usage of integrases in microbial engineering, biosynthesis, and engineering of tryptophan derived metabolites, regulation and discovery of fungal natural products, and elucidation and localization of plant pathways.

Preface
Chapter One: Comparative Biochemistry and In Vitro Pathway Reconstruction as Powerful Partners in Studies of Metabolic Diversity
- Abstract
- 1 Introduction
- 2 Characterization of Specialized Metabolite Diversity Using LC/MS and NMR
- 3 Phylogeny-Driven Analysis of the Biochemical Basis of Specialized Metabolite Diversity
- 4 Conclusions
- Acknowledgments
Chapter Two: De Novo Deep Transcriptome Analysis of Medicinal Plants for Gene Discovery in Biosynthesis of Plant Natural Products
- Abstract
- 1 Methods for Transcriptomic Study Prior to High-Throughput Sequencing
- 2 Deep Transcriptome Outline
- 3 Preparation of the Plant Materials and RNA Extraction
- 4 cDNA Library Construction
- 5 High-Throughput Sequencing
- 6 Data Interpretation
- 7 Application of RNA-Seq on Medicinal Plants and Perspectives
- Acknowledgments
Chapter Three: Genomics-Based Discovery of Plant Genes for Synthetic Biology of Terpenoid Fragrances: A Case Study in Sandalwood oil Biosynthesis
- Abstract
- 1 Introduction
- 2 Prior Knowledge of Sandalwood TPSs and P450s and Development of a Hypothesis
- 3 Replication, Sampling, and Statistical Design
- 4 Defining Temporal and Spatial Variables for Tissue Sampling
- 5 Tissue Sampling
- 6 Metabolite Profiling
- 7 Isolation of High-Quality RNA from Recalcitrant Tissues
- 8 Transcriptome Sequencing and De Novo Assembly
- 9 Transcriptome Mining and Annotation
- 10 Expression Analysis and Candidate Gene Selection
- 11 Functional Characterization of Candidate Genes
- 12 Product Identification
- Acknowledgments
Chapter Four: A Workflow for Studying Specialized Metabolism in Nonmodel Eukaryotic Organisms
- Abstract
- 1 Introduction
- 2 “Omics”-Based Novel Specialized Metabolic Pathway Discovery
- 3 Structure–Function Analysis of Specialized Metabolic Enzymes
- 4 Reconstitution of Specialized Metabolic Pathways in Heterologous Systems
- 5 Summary
- Acknowledgments
Chapter Five: Gene Discovery for Synthetic Biology: Exploring the Novel Natural Product Biosynthetic Capacity of Eukaryotic Microalgae
- Abstract
- 1 Introduction
- 2 Natural Product Synthases
- 3 Genome Mining for the Identification of Natural Products
- 4 Natural Product Discovery
- 5 Conclusions
- Acknowledgments
Chapter Six: cis-Prenyltransferase and Polymer Analysis from a Natural Rubber Perspective
- Abstract
- 1 Introduction
- 2 Rationale: Observation of Revertants from rer2 Mutant
- 3 Generation of rer2 and srt1 Double Knockout Yeast Strain
- 4 Complementation of rer2Δ srt1Δ with CPT and CBP
- 5 CPT Biochemical Assay Using Yeast Microsomes
- 6 General Discussion
- Acknowledgments
Chapter Seven: Generation and Functional Evaluation of Designer Monoterpene Synthases
- Abstract
- 1 Introduction
- 2 Equipment
- 3 Materials
- 4 Step 1—Generation of Expression Constructs
- 5 Step 2—Production of Purified, Recombinant Target Enzyme
- 6 Step 3—Functional Evaluation of Recombinant Monoterpene Synthases
- 7 Conclusions
- Acknowledgments
Chapter Eight: Prequels to Synthetic Biology: From Candidate Gene Identification and Validation to Enzyme Subcellular Localization in Plant and Yeast Cells
- Abstract
- 1 Introduction
- 2 Identification of Candidate Genes Through Transcriptomic Data Mining and Analysis
- 3 Validation of Candidate Gene Function by Biolistic-Mediated VIGS
- 4 Studying the Subcellular Localization of Biosynthetic Pathway Enzymes in Plant and Yeast Cells to Alleviate Bottlenecks in Bioengineering Approaches
- 5 Concluding Remarks
- Acknowledgments
Chapter Nine: Functional Expression and Characterization of Plant ABC Transporters in Xenopus laevis Oocytes for Transport Engineering Purposes
- Abstract
- 1 Introduction
- 2 Preparation of cDNA of Plant ABC Transporter Genes by In Planta “Exon Engineering”
- 3 ABC Transporter Expression in Xenopus Oocytes
- 4 Optimization of Transport Assay for Diffusible ABA in Xenopus Oocytes
- 5 Case Study: Characterization of the ABA Exporter at ABCG25 in Xenopus Oocytes
- 6 Conclusions
- Acknowledgments
Chapter Ten: Quantifying the Metabolites of the Methylerythritol 4-Phosphate (MEP) Pathway in Plants and Bacteria by Liquid Chromatography–Triple Quadrupole Mass Spectrometry
- Abstract
- 1 Introduction
- 2 Preparation of Stable Isotope-Labeled Internal Standards
- 3 Extraction of Methylerythritol Phosphate Pathway Intermediates from Biological Sources
- 4 Analysis of Methylerythritol Phosphate Pathway Metabolites by LC–MS/MS
- 5 Discussion and Summary
Chapter Eleven: Establishing the Architecture of Plant Gene Regulatory Networks
- Abstract
- 1 Introduction
- 2 The cis-Regulatory Apparatus
- 3 The Trans-Acting Factors
- 4 Transcription Factor Centered Approaches
- 5 Gene-Centered Approaches
- 6 Resources for Studying Plant GRNs
- 7 Conclusions
- Acknowledgment
Chapter Twelve: Engineering of Tomato Glandular Trichomes for the Production of Specialized Metabolites
- Abstract
- 1 Introduction
- 2 Materials and Technology
- 3 Proof of Concept: Targeted Expression of a Terpene Precursor Gene in Tomato Glandular Trichomes
- 4 Summary
- Acknowledgments
Chapter Thirteen: Tomato Fruits—A Platform for Metabolic Engineering of Terpenes
- Abstract
- 1 Introduction
- 2 Terpenoid Formation in Tomato Fruits
- 3 Transgene Expression in Ripening Tomato Fruits
- 4 Overexpression of Terpene Biosynthetic Genes in Tomato Fruits
- 5 Analysis of Terpenes in Tomato Fruits
- 6 Conclusions
- Acknowledgments
Chapter Fourteen: Libraries of Synthetic TALE-Activated Promoters: Methods and Applications
- Abstract
- 1 Introduction
- 2 Construction of Libraries of Synthetic Promoters Using Golden Gate Cloning
- 3 Analyzing Promoter Activity in Transient Assays
- 4 Conclusion
Author Index
Subject Index

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Synthetic Biology and Metabolic Engineering in Plants and Microbes Part B: Metabolism in Plants

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Sarah E O'Connor