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Tandem Metal-Oxide Affinity Chromatography for Enhanced Depth of Phosphoproteome Analysis

In eukaryotic cells many diverse cellular functions are regulated by reversible protein phosphorylation. In recent years, phosphoproteomics has become a powerful tool to study protein phosphorylation because it allows unbiased localization, and site-specific quantific ...

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Simultaneous Identification and Quantification of Nitrosylation Sites by Combination of Biotin Switch and ICAT Labeling

S-nitrosylation is a widespread modification of proteins. In plants, most information available to date regarding this modification was obtained using nitric oxide donors and concerned the proteins but not the identification of cysteine residues specifically modified in the pr ...

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StageTip-Based HAMMOC, an Efficient and Inexpensive Phosphopeptide Enrichment Method for Plant Shotgun Phosphoproteomics

Phosphopeptide enrichment is the most critical step for successful LC-MS/MS-based shotgun phosphoproteomics. Recent technological improvements have made selective phosphopeptide enrichment from non-fractionated whole cell lysate digests with a single-step proce ...

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Protein Extraction and Gel-Based Separation Methods to Analyze Responses to Pathogens in Carnation (Dianthus caryophyllus L)

We are currently using a 2-DE-based proteomics approach to study plant responses to pathogenic fungi by using the carnation (Dianthus caryophyllus L)–Fusarium oxysporum f. sp. dianthi pathosystem. It is clear that the protocols for the first stages of a standard proteomics workflow must be ...

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Differential Proteome and Secretome Analysis During RicePathogen Interaction

Substantial evidences implicate that sample preparation and protein extraction in proteomic studies of plant–pathogen interactions are critical to understand cross talk between host and pathogen. Therefore, interest is growing in applying proteomics techniques to inves ...

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Proteomics of Field Samples in Radioactive Chernobyl Area

Two serious nuclear accidents during the past quarter of a century contaminated large agricultural areas with radioactivity. The remediation and possible recovery of radio-contaminated areas for agricultural purposes require comprehensive characterization of plants g ...

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Isolation of Leaf Peroxisomes from Arabidopsis for Organelle Proteome Analyses

The isolation of cell organelles from model organisms in high purity is important for biochemical analyses of single proteins, entire metabolic pathways, and protein complexes and is absolutely essential for organelle proteome analyses. The efficient enrichment of nearly all cell ...

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Separation of the Plant Golgi Apparatus and Endoplasmic Reticulum by Free-Flow Electrophoresis

Free-flow electrophoresis (FFE) is a technique for separation of proteins, peptides, organelles, and cells. With zone electrophoresis (ZE-FFE), organelles are separated according to surface charge. The plant Golgi and endoplasmic reticulum (ER) are similar in density and are there ...

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Plant Mitochondrial Proteomics

Mitochondria are responsible for a number of major biochemical processes in plant cells including oxidative phosphorylation and photorespiration. Traditionally their primary role has been viewed as the oxidation of organic acids via the tricarboxylic acid cycle and the synthe ...

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Shotgun Proteomics of Plant Plasma Membrane and Microdomain Proteins Using Nano-LC-MS/MS

Shotgun proteomics allows the comprehensive analysis of proteins extracted from plant cells, subcellular organelles, and membranes. Previously, two-dimensional gel electrophoresis-based proteomics was used for mass spectrometric analysis of plasma membrane protei ...

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Quantitative Proteomic Analysis of Intact Plastids

Plastids are specialized cell organelles in plant cells that are differentiated into various forms including chloroplasts, chromoplasts, and amyloplasts, and fulfill important functions in maintaining the overall cell metabolism and sensing environmental factors such as ...

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Isolation of the Plant Cytosolic Fraction for Proteomic Analysis

The cytosol is the fluid portion of the cell that is not partitioned by membranes. It contains a highly diverse collection of substances and is central to many essential cellular processes ranging from signal transduction, metabolite production and transport, protein biosynthesis and ...

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Suspension-Cultured Plant Cells as a Tool to Analyze the Extracellular Proteome

Suspension-cultured cells (SCC) are generally considered the most suitable cell systems to carry out scientific studies, including the extracellular proteome (secretome). SCC are initiated by transferring friable callus fragments into flasks containing liquid culture me ...

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Pollen Cultivation and Preparation for Proteomic Studies

The quality of the collected experimental data very much depends on the quality of the biological starting material. Especially the proteome analysis of a highly dynamic system like the germinating and tube-growing pollen grain needs several precautions which allow an accurate and acc ...

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Back to Osborne. Sequential Protein Extraction and LC-MS Analysis for the Characterization of the Holm Oak Seed Proteome

It is impossible to capture in just one experiment all or most of the total set of protein species that constitute the cell’s proteome. Thus, according to our results, and even considering that they depend on the experimental system carried out (plant, yeast, fungi, or bacteria), the best protein extr ...

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Xylem Sap Proteomics

Proteomic analysis of xylem sap has recently become a major field of interest to understand several biological questions related to plant development and responses to environmental clues. The xylem sap appears as a dynamic fluid undergoing changes in its proteome upon abiotic and biotic ...

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Seed Proteomics

Rather than providing a single specific protocol, the inclusive area of seed proteomics is reviewed; methods are described and compared and primary literature citations are provided. The limitations and challenges of proteomics as an approach to study seed biology are emphasized. The p ...

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An Improved Detergent-Compatible Gel-Fractionation LC-LTQ-Orbitrap-MS Workflow for Plant and Microbial Proteomics

In proteomics, liquid chromatography coupled to mass spectrometry (LC-MS/MS) is an invaluable technique to accurately identify and quantify large sets of proteins. In this chapter we show a time-effective, and detergent compatible, Ge-LC-LTQ-Orbitrap/MS proteomics workflow. T ...

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Proteome Analysis of Orphan Plant Species, Fact or Fiction

Biological research has focused in the past on model organisms and most of the functional genomics studies in the field of plant sciences are still performed on model species or reference species that are characterized to a great extent. However, numerous non-model plants are essential as food, ...

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Medicago truncatula Proteomics for Systems Biology: Novel Rapid Shotgun LC-MS Approach for Relative Quantification Based on Full-Scan Selective Peptid

Medicago truncatula has become the focus of systems biology research for improved legume crop breeding. In plant systems biology, several comparative studies have been carried out using liquid chromatography shotgun mass spectrometry (LC-MS/MS) and database-dependent prote ...

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