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Two-Dimensional Electrophoresis for Plant Proteomics

  • Protocol
Plant Proteomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 355))

Abstract

Two-dimensional gel electrophoresis (2-DE) with immobilized pH gradients (IPGs) combined with protein identification by mass spectrometry (MS) is currently the workhorse for proteome analysis. 2-DE allows separation of highly complex mixtures of proteins according to isoelectric point (pI), molecular mass (M r), solubility, and relative abundance and delivers a map of intact proteins, which reflects changes in protein expression level, isoforms, or posttranslational modifications. 2-DE can resolve more than 5000 proteins simultaneously (approx 2000 proteins routinely) and can detect and quantify <1 ng of protein per spot. Today’s 2-DE technology with IPGs has overcome the former limitations of carrier ampholyte-based 2-DE with respect to reproducibility, handling, resolution, and separation of very acidic and/or basic proteins. The development of IPGs between pH 2.5 and 12 has allowed the analysis of very alkaline proteins and the construction of the corresponding databases. Narrow pH range IPGs provide increased resolution (ΔpI = 0.001) and, in combination with prefractionation methods, permit the detection of low abundance proteins. In this article we provide a comprehensive protocol of the current 2-DE technology for plant proteome analysis and describe in detail the individual steps of this technique.

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Author information

Authors and Affiliations

  1. Fachgebiet Proteomik, Am Forum 2, Technische Universität München, Freising-Weihenstephan, Germany

    Walter Weiss & Angelika Görg

Authors
  1. Walter Weiss
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  2. Angelika Görg
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Editor information

Editors and Affiliations

  1. UMR de Génétique Végétale, La Ferme du Moulon, Gif-sur-Yvette, France

    Hervé Thiellement , Michel Zivy  & Catherine Damerval ,  & 

  2. UMR de Chimie Biologique, Thiverval Grignon, France

    Valérie Méchin

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© 2007 Humana Press Inc., Totowa, NJ

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Weiss, W., Görg, A. (2007). Two-Dimensional Electrophoresis for Plant Proteomics. In: Thiellement, H., Zivy, M., Damerval, C., Méchin, V. (eds) Plant Proteomics. Methods in Molecular Biology, vol 355. Humana Press. https://doi.org/10.1385/1-59745-227-0:121

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  • DOI: https://doi.org/10.1385/1-59745-227-0:121

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-635-1

  • Online ISBN: 978-1-59745-227-4

  • eBook Packages: Springer Protocols

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