Research Article

Protein extraction for two-dimensional electrophoresis from olive leaf, a plant tissue containing high levels of interfering compounds

  1. Wei Wang1,*,
  2. Monica Scali1,
  3. Rita Vignani1,
  4. Antonia Spadafora2,
  5. Elisabetta Sensi1,
  6. Silvia Mazzuca2,
  7. Mauro Cresti1

Article first published online: 16 JUL 2003

DOI: 10.1002/elps.200305500

Issue

ELECTROPHORESIS

ELECTROPHORESIS

Volume 24, Issue 14, pages 2369–2375, No. 14 July 2003

Additional Information

How to Cite

Wang, W., Scali, M., Vignani, R., Spadafora, A., Sensi, E., Mazzuca, S. and Cresti, M. (2003), Protein extraction for two-dimensional electrophoresis from olive leaf, a plant tissue containing high levels of interfering compounds. ELECTROPHORESIS, 24: 2369–2375. doi: 10.1002/elps.200305500

Author Information

  1. 1

    Dipartimento di Scienze Ambientali, University of Siena, Siena, Italy

  2. 2

    Laboratorio di Citofisiologia Vegetale, Dipartimento di Ecologia, University of Calabria, Rende, Italy

*Dipartimento di Scienze Ambientali, University of Siena, Via P A Mattioli 4, I-53100 Siena, Italy Fax: +39-0577-232860

Publication History

  1. Issue published online: 16 JUL 2003
  2. Article first published online: 16 JUL 2003
  3. Manuscript Received: 11 MAR 2003

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Keywords:

  • Contaminants;
  • Electrophoresis;
  • Immunoblot;
  • Olive leaf;
  • Plant protein extraction

Abstract

The purpose of this research is to establish a routine procedure for the application of proteomic analysis to olive tree. Olive leaf tissue is notoriously recalcitrant to common protein extraction methods due to high levels of interfering compounds. We developed a protocol for isolating proteins suitable for two-dimensional electrophoresis (2-DE) from olive leaf. The remarkable characteristics of the protocol include: (i) additional grinding dry acetone powder of leaf tissue to a finer extent, (ii) after extensive organic solvent washes to remove pigments, lipids etc., using aqueous tricholoroacetic acid washes to remove water-soluble contaminants, and (iii) phenol extraction of proteins in the presence of sodium dodecyl sulfate. The final protein preparation is free of interfering compounds based on its well-resolved 2-DE patterns. The protocol can be completed within 3 h, and protein yield is approximately 2.49 mg⋅g−1 of aged leaf. We also evaluated the protocol by immunoblotting with anti-tyrosinate α-tubulin antibody. To our knowledge, this is the first time that a protocol for protein extraction from olive leaf appears to give satisfactory and reproducible results. The protocol is expected to be applicable to other recalcitrant plant tissues and could be of interest to laboratories involved in plant proteomics.

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