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Applications of Difference Gel Electrophoresis (DIGE) in the Study of Microorganisms

  • Kathleen Trautwein
  • Ralf Rabus
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1841)

Abstract

OMICs-based investigations of microorganisms are becoming more and more widespread in the upcoming era of systems and synthetic biology. Here, proteomics plays a key role and two-dimensional difference gel electrophoresis (2D DIGE) remains the “gold-standard” for globally determining protein abundance changes on a quantitative and statistically confident level—in particular also for laboratories not having full-cycle proteomic facilities at their disposal. In this contribution we summarize our methodological procedures and experiences with 2D DIGE accumulated over the past 15 years.

Key words

2D DIGE 2DE Fluorescence labeling Image analysis Quantitative proteome profiling Differential proteomics Microorganisms 

Notes

Acknowledgment

Proteomic research (incl. DIGE) in our laboratory was supported by the German Federal Ministry of Education and Research BMBF (REGX-project), the Max Planck Society, and the Deutsche Forschungsgemeinschaft (SPP 1319, SFB TRR 51, GRK 1885).

References

  1. 1.
    Wöhlbrand L, Trautwein K, Rabus R (2013) Proteomic tools for environmental microbiology—a roadmap from sample preparation to protein identification and quantification. Proteomics 13(18–19):2700–2730PubMedGoogle Scholar
  2. 2.
    Rabus R, Trautwein K, Wöhlbrand L (2014) Towards habitat-oriented systems biology of “Aromatoleum aromaticum” EbN1: chemical sensing, catabolic network modulation and growth control in anaerobic aromatic compound degradation. Appl Microbiol Biotechnol 98(8):3371–3388CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Rabus R (2014) Fifteen years of physiological proteo(geno)mics with (marine) environmental bacteria. Arch Physiol Biochem 120(5):173–187CrossRefPubMedGoogle Scholar
  4. 4.
    Rabilloud T, Chevallet M, Luche S, Lelong C (2010) Two-dimensional gel electrophoresis in proteomics: past, present and future. J Proteome 73(11):2064–2077CrossRefGoogle Scholar
  5. 5.
    Görg A, Obermaier C, Boguth G, Harder A, Scheibe B, Wildgruber R, Weiss W (2000) The current state of two-dimensional electrophoresis with immobilized pH gradients. Electrophoresis 21(6):1037–1053CrossRefPubMedGoogle Scholar
  6. 6.
    Görg A, Weiss W, Dunn MJ (2004) Current two-dimensional electrophoresis technology for proteomics. Proteomics 4(12):3665–3685CrossRefPubMedGoogle Scholar
  7. 7.
    Hoving S, Voshol H, van Oostrum J (2000) Towards high performance two-dimensional gel electrophoresis using ultrazoom gels. Electrophoresis 21(13):2617–2621CrossRefPubMedGoogle Scholar
  8. 8.
    Ünlü M, Morgan ME, Minden JS (1997) Difference gel electrophoresis: a single gel method for detecting changes in protein extracts. Electrophoresis 18(11):2071–2077CrossRefPubMedGoogle Scholar
  9. 9.
    Gade D, Thiermann J, Markowsky D, Rabus R (2003) Evaluation of two-dimensional difference gel electrophoresis for protein profiling. Soluble proteins of the marine bacterium Pirellula sp. strain 1. J Mol Microbiol Biotechnol 5(4):240–251CrossRefPubMedGoogle Scholar
  10. 10.
    Rabus R, Trautwein K (2010) Proteogenomics to study the anaerobic degradation of aromatic compounds and hydrocarbons. In: Timmis KN (ed) Handbook of hydrocarbon and lipid microbiology. Springer, Berlin, pp 4385–4405CrossRefGoogle Scholar
  11. 11.
    Neuhoff V, Arold N, Taube D, Ehrhardt W (1988) Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250. Electrophoresis 9(6):255–262CrossRefPubMedGoogle Scholar
  12. 12.
    Strijkstra A, Trautwein K, Roesler S, Feenders C, Danzer D, Riemenschneider U, Blasius B, Rabus R (2016) High performance CCD camera system for digitalization of 2D DIGE gels. Proteomics 16(14):1975–1979CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Zech H, Echtermeyer C, Wöhlbrand L, Blasius B, Rabus R (2011) Biological versus technical variability in 2-D DIGE experiments with environmental bacteria. Proteomics 11(16):3380–3389CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institute for Chemistry and Biology of the Marine Environment (ICBM)Carl von Ossietzky University OldenburgOldenburgGermany

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