Xylem: Methods and Protocols (Methods in Molecular Biology, 2722)

✍ Scribed by Javier Agusti (editor)

• Publisher: Humana
• Year: 2023
• Tongue: English
• Leaves: 244
• Edition: 2
• Category: Library

✦ Synopsis

This second edition provides detailed techniques used for the study and characterization of the plant vascular system, with a central focus on the xylem tissue. Chapters are organized in three main sections covering; analysis of xylem development, xylem characterization though imaging techniques, and analysis of the xylem composition. Written in the highly 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 laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls.

Authoritative and cutting-edge, Xylem: Methods and Protocols, Second Edition aims to be a useful and practical guide to new researchers and experts looking to expand their knowledge.

✦ Table of Contents

Preface
Part I: Xylem Transport, Functional Dynamics and Modelling (Chaps. 1, 2, 3, and 4)
Part II: Xylem Development and Evolution (Chaps. 5, 6, and 7)
Part III: Xylem Diseases (Chaps. 8 and 9)
Part IV: Xylem Composition and Imaging (Chaps. 10, 11, 12, 13, 14, and 15)
Contents
Contributors
Part I: Xylem Transport, Functional Dynamics and Modelling
Chapter 1: Monitoring Xylem Transport in Arabidopsis thaliana Seedlings Using Fluorescent Dyes
1 Introduction
2 Materials
2.1 Applying Fluorescent Dye Solutions to Arabidopsis Seedling Roots
2.2 Monitoring Xylem Root-to-Shoot Transport
2.3 Monitoring Xylem-Transported Fluorescent Dyes on the Cellular Level
3 Methods
3.1 Applying Fluorescent Dyes to Arabidopsis Seedlings
3.2 Monitoring Xylem Root-to-Shoot Transport
3.3 Monitoring Xylem Transported CFDA on the Cellular Level
4 Notes
References
Chapter 2: Modeling and Analyzing Xylem Vulnerability to Embolism as an Epidemic Process
1 Relevance of Xylem Embolism and Its Quantification
2 Vulnerability Curves and Functions Suited for Fitting Them
3 Mechanistic Approaches to Describe Xylem Vulnerability for Embolism
3.1 Modeling Embolism by Graphs and Network Theory
3.2 Modeling Embolism by Epidemic Theory
3.2.1 The Classic SIR Model
3.2.2 Application of the SIR Model to Xylem Embolism and Air Seeding
3.2.3 Differences and Links Between the SIR Approach and Network Models-the Relevance and Role of Spatial Structure and Probab...
4 Conclusions
References
Chapter 3: Modeling Xylem Functionality Aspects
1 Introduction
1.1 Previous Approaches to Describe Xylem Functionality
1.2 The Plant Hydraulic System
1.3 Plant Architecture
2 Materials and Methods
2.1 Governing Equations and Physical Laws
2.1.1 Conservation of Energy
2.1.2 Conservation of Momentum
2.1.3 Conservation of Mass
2.2 Implementation Framework
2.2.1 Representing and Encoding Plant Architecture
2.2.2 Functional-Structural Plant Modeling
2.2.3 Numerical Solvers
2.2.4 Auxiliary Models
Radiation
Stomatal Conductance and Photosynthesis
3 Notes
References
Chapter 4: Detecting and Quantifying Xylem Embolism by Synchrotron-Based X-Ray Micro-CT
1 Introduction
2 Materials
2.1 Synchrotron-Based Micro-CT Facility
2.2 Plant Material
2.3 Sample Preparation
2.4 Additional Measurements
2.5 Reconstruction Software
2.6 Image Analysis Software for Xylem Embolism Quantification
3 Methods
3.1 Technical Specification
3.2 How to Prepare Plant Samples
3.3 Acquisition Procedure
3.4 Additional Measurements
3.5 Image Reconstruction
3.6 Image Analysis
3.7 Xylem Vulnerability Curve (VC) Estimation
4 Notes
References
Part II: Xylem Development and Evolution
Chapter 5: Analysis of Xylem Cells by Nucleus-Based Transcriptomics and Chromatin Profiling
1 Introduction
2 Material
2.1 Plant Material
2.2 Nucleus Isolation Using a Centrifuge
2.3 Nucleus Isolation Without Using a Centrifuge
2.4 Nucleus Sorting
2.5 RNA Extraction and SMART-seq2 Amplification
2.6 Tagmentation for ATAC-seq Analysis
3 Methods
3.1 Nucleus Isolation Using Centrifuges
3.2 Nucleus Isolation Without Using Centrifuges (See Note 4)
3.3 Nucleus Sorting Using Cell Sorters
3.4 RNA Extraction, Smart-seq2 Amplification and Purification (See Note 6)
3.5 Tagmentation for ATAC-seq Analysis
4 Notes
References
Chapter 6: Quantification of Xylem-Specific Thermospermine-Dependent Translation of SACL Transcripts with Dual Luciferase Repo...
1 Introduction
2 Materials
2.1 Agroinfiltration of N. benthamiana Leaves and Tspm Treatment
2.2 Protoplast PEG-Calcium Transfection
2.3 Luciferase Assay
3 Methods
3.1 Agroinfiltration of N. benthamiana Leaves and Tspm Treatments
3.2 Protoplast PEG-Calcium Transfection and Tspm Treatments (This Protocol Has Been Adapted from)
3.3 Dual Glo Luciferase Assay
4 Notes
References
Chapter 7: Fossil Wood Analyses: Several Examples from Five Case Studies in the Area of Central and NW Bohemia, Czech Republic
1 Introduction
2 Paleozoic Woods
2.1 Case Study 1: Kladno-Rakovník Basin
2.1.1 Completeness of Fossil Record
2.1.2 Unifacial vs. Bifacial Cambium
2.1.3 Influence of Environment on Mode of Preservation
3 Mesozoic Woods
3.1 Case Study 2: Bohemian Cretaceous Basin
3.1.1 Formation of Tyloses and Its Significance
3.1.2 Stem vs. Crown Group
3.1.3 Wide Concept of Fossil Wood Genera
4 Cenozoic Woods
4.1 Case Study 3: Most Basin
4.1.1 Influence of Preservation on Wood Anatomy and Preservation Potential
4.1.2 Mosaic´´ Species 4.1.3 Early vs. Late Wood 4.2 Case Study 4: České stredohorí Mountains 4.2.1 Discrepancy Between the Record of Wood and Other Organs 4.2.2 Unambiguity of Scientific Terminology 4.2.3 Stem vs. Root Wood 4.3 Case Study 5: Doupovské Hory Mountains 4.3.1 Wood of Extinct Plants 4.3.2 Definition ofWood Type´´
4.3.3 Subjective vs. Objective Methods of Palaeoclimatic Reconstruction
5 Conclusions
References
Part III: Xylem Diseases
Chapter 8: Isolation and Reproductive Structures Induction of Fungal Pathogens Associated with Xylem and Wood Necrosis in Grap...
1 Introduction
2 Materials
2.1 Preparation of Culture Media
2.2 Plant Material Preparation and Fungal Isolation
3 Methods
3.1 Preparation of Culture Media for Fungal Isolation
3.2 Fungal Isolation from Trunk Samples
3.3 Fungal Isolation from Root Samples
3.4 Fruiting Bodies (Pycnidia) Induction in Specific Culture Media
4 Notes
References
Chapter 9: Determination of De Novo Suberin-Lignin Ferulate Deposition in Xylem Tissue Upon Vascular Pathogen Attack
1 Introduction
2 Materials
2.1 Plant Varieties and Plant Growth Materials
2.2 Bacterial Strains and Bacterial Culture
2.3 Tissue Sectioning
2.4 Histological Materials
3 Methods
3.1 Bacterial Inoculation in Plants (Soil-Drenching Method)
3.2 Histochemical Analysis
3.3 Lignin Staining
3.4 Detecting Ferulate Deposition
3.5 Detecting Suberin Aliphatics
3.6 Deciphering the Composition and Structure of the Cell Wall-Deposited Compounds
3.6.1 Solvent Extraction for Removing Nonstructural Plant Cell Wall Components
3.6.2 Ball-Milling
3.6.3 Lignin/Suberin Isolation by Enzymatic Removal of Polysaccharides
3.6.4 2D-NMR Analysis
3.6.5 Assignation and Quantitation of 2D-HSQC Correlation Signals
4 Notes
References
Part IV: Xylem Composition and Imaging
Chapter 10: Quantification of Tracheary Elements Types in Mature Hypocotyl of Arabidopsis thaliana
1 Introduction
2 Materials
3 Methods
3.1 Vegetal Material
3.2 Maceration (See Note 3)
3.3 Microscope Visualization
3.4 Xylem Cell Types
4 Notes
References
Chapter 11: Histochemical Detection of Peroxidase and Laccase Activities in Populus Secondary Xylem
1 Introduction
2 Materials
2.1 General Remarks
2.2 Equipment
2.3 Reagents
2.4 Detection of Xylem Cell Wall PRXs and LACs
2.5 Enzyme Inhibition and ROS Scavengers
2.6 Monitoring ROS
3 Methods
3.1 Detection of Xylem Cell Wall PRXs and LACs
3.2 Enzyme Inhibition and ROS Scavengers
3.3 Monitoring ROS
4 Notes
References
Chapter 12: Lignin Analysis by HPLC and FTIR: Spectra Deconvolution and S/G Ratio Determination
1 Introduction
2 Materials
2.1 Sampling and Extractive-Free Wood Isolation
2.2 Klason Lignin
2.3 FTIR Spectroscopy
2.4 Alkaline Nitrobenzene Oxidation of Lignin
2.5 Determination of the S/G Ratio by HPLC
3 Methods
3.1 Sample Preparation
3.2 Obtaining Extractive-Free Wood
3.3 Klason Lignin
3.4 Preparation of 1% KBr Pellets for FTIR Spectroscopy
3.5 Alkaline Nitrobenzene Oxidation and S/G Ratio Determination by HPLC
3.6 Deconvolution of FTIR Spectra for the Determination of the S/G Ratio
4 Notes
References
Chapter 13: Inducible Pluripotent Suspension Cell Cultures (iPSCs) to Study Plant Cell Differentiation
1 Introduction
2 Materials
2.1 Equipment and Infrastructure
2.2 Cell Culture Media, Hormones, and Solutions
3 Methods
3.1 Initiating Cell Suspension Culture from Calli
3.2 Screening of Hormone-Habituated Growth in Cell Suspension Cultures to Establish iPSCs
3.3 Stable Genetic Transformation of Habituated Cell Suspensions
3.4 Triggering the Differentiation of iPSCs into Specific Cell Types
3.5 Monitoring Cell Culture Growth, Viability, and Differentiation
3.6 Real-Time Live Cell Imaging of Cell Differentiation in iPSCs
3.7 Drug-Induced Modulation of Cell Differentiation or Uncoupling of Distinct Specialization Stage
3.8 Extracting Biomolecules During Cell Differentiation Time-Course
4 Notes
References
Chapter 14: Bulk and In Situ Quantification of Coniferaldehyde Residues in Lignin
1 Introduction
2 Materials
2.1 Equipment and Infrastructure
2.2 Chemical Solutions
3 Methods
3.1 Producing Synthetic Lignin with Only Coniferaldehyde Units
3.2 Total Coniferaldehyde Unit Quantification in Lignin Using Pyrolysis-GC/MS
3.3 Terminal and Internal Ether-Linked Coniferaldehyde Unit Quantification in Lignin Using Thioacidolysis-GC/MS
3.4 Total Coniferaldehyde Unit Quantification in Lignin In Situ Using the Wiesner Test
3.5 Terminal Coniferaldehyde Unit Quantification in Lignin In Situ Using Raman Spectroscopy
4 Notes
References
Chapter 15: Clearing of Vascular Tissue in Arabidopsis thaliana for Reporter Analysis of Gene Expression
1 Introduction
2 Materials
2.1 Plant Lines and Growth Conditions
2.2 Solutions
2.2.1 GUS Stock Solutions
2.2.2 Fixing Solution for Imaging Fluorescent Protein Tags
2.2.3 Clearing Solution
2.2.4 Staining Solutions
2.3 Materials for Sample Processing and Microscopy
3 Methods
3.1 Plant Cultivation In Vitro
3.2 Plant Cultivation on Soil
3.3 Seedlings GUS Staining and Clearing
3.4 Hypocotyls Clearing for FP Visualization Combined with Cell Wall Staining
3.5 Imaging
3.5.1 Imaging of Cleared GUS Stained Seedlings
3.5.2 Imaging of FPs Combined with Cell Wall Staining in Cleared Hypocotyl Sections
4 Notes
References
Index