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    Protocols in Current Issue
    Apoplastic Expression of CARD1-ecto Domain in Nicotiana benthamiana and Purification from the Apoplastic Fluids
    Authors:  Nobuaki Ishihama, Anuphon Laohavisit, Kaori Takizawa and Ken Shirasu, date: 04/20/2022, view: 769, Q&A: 0
    [Abstract]

    The protein expression and purification process is an essential initial step for biochemical analysis of a protein of interest. Traditionally, heterologous protein expression systems (such as E. coli, yeast, insect cells, and cell-free) are employed for plant protein expression, although a plant expression system is often desirable for plant

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    Split-luciferase Complementation Imaging Assay to Study Protein-protein Interactions in Nicotiana benthamiana
    Authors:  Liping Wang, Gang Yu, Alberto P. Macho and Rosa Lozano-Durán, date: 12/05/2021, view: 1835, Q&A: 0
    [Abstract]

    The experimental identification of protein-protein interactions (PPIs) is critical to understand protein function. Thus, a plethora of sensitive and versatile approaches have been developed to detect PPIs in vitro or in vivo, such as protein pull-down, yeast two-hybrid (Y2H), co-immunoprecipitation (co-IP), and bimolecular fluorescence

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    Visualization and Quantification of Cell-to-cell Movement of Proteins in Nicotiana benthamiana
    Authors:  Mila C Blekemolen, Mariliis Tark-Dame and Frank L.W Takken, date: 12/20/2018, view: 5328, Q&A: 0
    [Abstract] Cell-to-cell movement of proteins through plasmodesmata is a widely-established mechanism for intercellular signaling in plants. Current techniques to study intercellular protein translocation rely on single-cell transformation using particle bombardment or transgenic lines expressing photo-inducible fluorophores. The method presented here allows ...
    Ubiquitin Proteasome Activity Measurement in Total Plant Extracts
    Authors:  Suayib Üstün and Frederik Börnke, date: 09/05/2017, view: 7551, Q&A: 0
    [Abstract] The fine-tuned balance of protein level, conformation and location within the cell is vital for the dynamic changes required for a cell to respond to a given stimulus. This requires the regulated turnover of damaged or short-lived proteins through the ubiquitin proteasome system (UPS). Thus, the protease activity of the proteasome is adjusted to ...
    Expression and Partial Purification of His-tagged Proteins in a Plant System
    Author:  Amancio de Souza, date: 09/05/2015, view: 11814, Q&A: 0
    [Abstract] Plant protein expression can be a challenging enterprise in any biochemical or molecular biology research project. Several heterologous systems like bacteria, yeast, insect cells and cell free systems have been used to produce plant proteins for in vitro experiments and structural characterization. However, due to particularities of plant ...
    Protein Immunoprecipitation Using Nicotiana benthamiana Transient Expression System
    Authors:  Fang Xu, Charles Copeland and Xin Li, date: 07/05/2015, view: 19350, Q&A: 0
    [Abstract] Nicotiana benthamiana (N. benthamiana) is a useful model system to transiently express protein at high level. This protocol describes in detail how to transiently express protein in N. benthamiana and how to carry out protein immunoprecipitation in this expression system. This protocol can be broadly used for ...
    Localization and Topology of Thylakoid Membrane Proteins in Land Plants
    Authors:  Salar Torabi, Magdalena Plöchinger and Jörg Meurer, date: 12/20/2014, view: 11385, Q&A: 0
    [Abstract] Thylakoids are a formation of flattened membrane vesicles and protein complexes found in cyanobacteria, algae and plants. In the chloroplasts of land plants the thylakoid membrane systems form a network of densely packed stacks called grana lamellae, which are connected by unstacked stroma lamellae. Photosystem II is mainly localized in the ...
    Fluorescence Recovery after Photobleaching (FRAP) Assay to Measure the Dynamics of Fluorescence Tagged Proteins in Endoplasmic Reticulum Membranes of Plant Cells
    Authors:  José Antonio Navarro, Marta Serra-Soriano and Vicente Pallás, date: 10/20/2014, view: 16585, Q&A: 0
    [Abstract] In this protocol, we used fluorescence recovery after photobleaching (FRAP) to measure the influence that some mutations and drug treatment have on mobility of a green fluorescent protein (GFP)-fused viral transmembrane protein into endoplasmic reticulum membranes (Serra-Soriano et al., 2014). The proteins of interest were transiently ...
    Extraction of Chloroplast Proteins from Transiently Transformed Nicotiana benthamiana Leaves
    Author:  Joern Klinkenberg, date: 09/20/2014, view: 17545, Q&A: 0
    [Abstract] This rapid protocol allows the extraction of chloroplast enriched proteins from Nicotiana benthamiana (N. benthamiana) leaves that were transiently transformed to express an epitope tagged protein of interest. Thus, it can serve as a tool to study the chloroplastidic localization of the protein of interest when it is combined ...
    SGR-based Reporter to Assay Plant Transcription Factor-promoter Interactions
    Authors:  Shisong Ma and Savithramma Dinesh-Kumar, date: 08/20/2014, view: 9512, Q&A: 0
    [Abstract] We developed an in vivo method to assay plant transcription factor (TF)–promoter interactions using the transient expression system in Nicotiana benthamiana (N. benthamiana) plants. The system uses the Arabidopsis stay green (SGR) gene as a reporter. Induction of SGR expression in N. ...



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