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Comparative Analyses of Protein Complexes by Blue Native DIGE

  • Katrin Peters
  • Hans-Peter BraunEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 854)

Abstract

Classically, DIGE is carried out on the basis of two-dimensional (2D) IEF/SDS PAGE. This allows comparative analyses of large protein sets. However, 2D IEF/SDS PAGE only poorly resolves hydrophobic proteins and is not compatible with native protein characterizations. Blue native PAGE represents a powerful alternative. Combined with CyDye labeling, blue native DIGE offers several useful applications like quantitative comparison of protein complexes of related protein fractions. Here we present a protocol for fluorophore labeling of native protein fractions for separation by blue native PAGE.

Key words

Blue native PAGE DIGE labeling Protein complexes Membrane proteins Mitochondria 

Abbreviations

2D

Two-dimensional

BN PAGE

Blue native polyacrylamide gel electrophoresis

BN/SDS PAGE

Blue native/sodium dodecyl sulfate polyacrylamide gel electrophoresis

DIGE

Difference gel electrophoresis

DMF

Dimethylformamide

IEF/SDS PAGE

Isoelectric focusing/sodium dodecyl sulfate polyacrylamide gel electrophoresis

pI

Isoelectric point

PMSF

Phenylmethylsulfonyl fluoride

Notes

Acknowledgments

The authors would like to thank Christina Rode, Institute for Plant Genetics, Leibniz Universität Hannover, for critical reading of the manuscript.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Institute for Plant GeneticsLeibniz Universität HannoverHannoverGermany

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