As scientist begin to understand the complexity of lipid signaling and its roles in plant biology, there is an increasing interest in their analysis. Due to the low abundancy and transient nature of some of these hydrophobic compounds, this is not always easy. In Plant Lipid Signaling Protocols, expert researchers in the field detail experimental approaches by which plant signaling lipids can be studied. These methods and techniques include analysis of plant signaling lipids, including detailed protocols to detect various relevant compounds by targeted or non-targeted approaches; to assay relevant enzyme activities in biological material or using recombinant enzymes; to test for specific binding of signaling lipids to protein partners; or to visualize signaling lipids or lipid-derived signals in living plant cells. 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 practical, Plant Lipid Signaling Protocols aids plant researchers in the continuing to study the roles of lipid signals.
This book presents timely and important tools needed to elucidate particular roles of phosphoinositides in plants. It includes easy-to-use, reproducible protocols and contains expert tips on troubleshooting and avoiding known pitfalls.
Analyzing Plant Signaling Phospholipids through 32Pi-Labeling and TLC.-
Analysis of D3-,4-,5-phosphorylated Phosphoinositides using HPLC.- Mass
Measurement of Polyphosphoinositides by Thin Layer- and Gas Chromatography.-
Measurement of Inositol (1,4,5) Trisphosphate in Plant Tissues by a
Competitive Receptor Binding Assay.- Quantification of Diacylglycerol by Mass
Spectrometry.- Distinguishing Phosphatidic Acid Pools from de novo Synthesis,
PLD and DGK.- Use of Phospholipase A2 for the Production of
Lysophospholipids.- Analysis and Quantification of Plant Membrane Lipids by
Thin-Layer Chromatography and Gas Chromatography.- Lipidomic Analysis of
Plant Membrane Lipids by Direct Infusion Tandem Mass Spectrometry.- Detection
and Quantification of Plant Sphingolipids by LC-MS.- Analysis of Defense
Signals in Arabidopsis thaliana Leaves by Ultra Performance Liquid
Chromatography/ Tandem Mass Spectrometry: Jasmonates, Salicylic Acid,
Abscisic Acid.- Analysis of Fatty Acid Amide Hydrolase (FAAH) Activity in
Plants.- Ionization Behavior of Polyphosphoinositides Determined via the
Preparation of pH Titration Curves using Solid State 31P NMR.-
Phosphatidylinositol Synthase Activity from Plant Membrane Fractions and from
E. coli-Expressed Recombinant Peptides.- Phosphatidylinositol 4-kinase and
Phosphatidylinositol 4-phosphate 5-kinase Assays.- Assaying Inositol and
Phosphoinositide Phosphatase Enzymes.- Determination of Phospholipase C
Activity In Vitro.- Assaying Non-Specific Phospholipase C (NPC) Activity.-
Assaying Different Types of Plant Phospholipase D Activities In Vitro.-
Measuring PLD Activity In Vivo.- Assaying Plant Phosphatidic Acid Phosphatase
Activity.- Assay of Phospholipase A Activity.- Lipid-Binding Analysis using a
Fat Blot Assay.- Liposome-binding Assays to Assess Specificity and Affinity
of Phospholipid-Protein Interactions.- Lipid affinity beads FromIdentifying
New Lipid Binding Proteins to Assessing their Binding Properties.- Using
Genetically Encoded Fluorescent Reporters to Image Lipid Signaling in Living
Plants.- Imaging Changes in Cytoplasmic Calcium using the Yellow Cameleon 3.6
Biosensor and Confocal Microscopy.- kinase Assays.- Assaying Inositol and
Phosphoinositide Phosphatase Enzymes.- Determination of Phospholipase C
Activity In Vitro.- Assaying Non-Specific Phospholipase C (NPC) Activity.-
Assaying Different Types of Plant Phospholipase D Activities In Vitro.-
Measuring PLD Activity In Vivo.- Assaying Plant Phosphatidic Acid Phosphatase
Activity.- Assay of Phospholipase A Activity.- Lipid-Binding Analysis using a
Fat Blot Assay.- Liposome-binding Assays to Assess Specificity and Affinity
of Phospholipid-Protein Interactions.- Lipid affinity beads From
Identifying New Lipid Binding Proteins to Assessing their Binding
Properties.- Using Genetically Encoded Fluorescent Reporters to Image Lipid
Signaling in Living Plants.- Imaging Changes in Cytoplasmic Calcium using the
Yellow Cameleon 3.6 Biosensor and Confocal Microscopy.
