Dataset Information

A possible approach for gel-based proteomic studies in recalcitrant woody plants.

ABSTRACT: Woody plants are particularly difficult to investigate due to high phenolic, resin, and tannin contents and laborious sample preparation. In particular, protein isolation from woody plants for two-dimensional gel electrophoresis (2-DE) is challenging as secondary metabolites negatively interfere with protein extraction and separation. In this study, three protein extraction protocols, using TCA, phenol and ethanol as precipitation or extraction agents, were tested in order to select the more efficient for woody recalcitrant plant gel-based proteomics. Grapevine leaves, pine needles and cork oak ectomycorrhizal roots were used to represent woody plant species and tissues. The phenol protocol produced higher quality 2-DE gels, with increased number of resolved spots, better spot focusing and representation of all molecular mass and isoelectric point ranges tested. In order to test the compatibility of the phenol extracted proteomes with protein identification several spots were excised from the phenol gels and analyzed by mass spectrometry (MALDI-TOF/TOF). Regardless the incomplete genome/protein databases for the plant species under analysis, 49 proteins were identified by Peptide Mass Fingerprint (PMF). Proteomic data have been deposited to the ProteomeXchange with identifier PXD000224. Our results demonstrate the complexity of protein extraction from woody plant tissues and the suitability of the phenol protocol for obtaining high quality protein extracts for efficient 2-DE separation and downstream applications such as protein identification by mass spectrometry.

SUBMITTER: Sebastiana M

PROVIDER: S-EPMC3663981 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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A possible approach for gel-based proteomic studies in recalcitrant woody plants.

Sebastiana Mónica M Figueiredo Andreia A Monteiro Filipa F Martins Joana J Franco Catarina C Coelho Ana Varela AV Vaz Fátima F Simões Tânia T Penque Deborah D Pais Maria Salomé MS Ferreira Sílvia S

SpringerPlus 20130508 1

Woody plants are particularly difficult to investigate due to high phenolic, resin, and tannin contents and laborious sample preparation. In particular, protein isolation from woody plants for two-dimensional gel electrophoresis (2-DE) is challenging as secondary metabolites negatively interfere with protein extraction and separation. In this study, three protein extraction protocols, using TCA, phenol and ethanol as precipitation or extraction agents, were tested in order to select the more eff ...[more]

PMID: 23724367

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Project description:Reactive oxygen species (ROS) are constantly produced by metabolically active plant cells. The concentration of ROS may determine their role, e.g., they may participate in signal transduction or cause oxidative damage to various cellular components. To ensure cellular homeostasis and minimize the negative effects of excess ROS, plant cells have evolved a complex antioxidant system, which includes ascorbic acid (AsA). AsA is a multifunctional metabolite with strong reducing properties that allows the neutralization of ROS and the reduction of molecules oxidized by ROS in cooperation with glutathione in the Foyer-Halliwell-Asada cycle. Antioxidant enzymes involved in AsA oxidation and reduction switches evolved uniquely in plants. Most experiments concerning the role of AsA have been performed on herbaceous plants. In addition to extending our understanding of this role in additional taxa, fundamental knowledge of the complex life cycle stages of woody plants, including their development and response to environmental factors, will enhance their breeding and amend their protection. Thus, the role of AsA in woody plants compared to that in nonwoody plants is the focus of this paper. The role of AsA in woody plants has been studied for nearly 20 years. Studies have demonstrated that AsA is important for the growth and development of woody plants. Substantial changes in AsA levels, as well as reduction and oxidation switches, have been reported in various physiological processes and transitions described mainly in leaves, fruits, buds, and seeds. Evidently, AsA exhibits a dual role in the photoprotection of the photosynthetic apparatus in woody plants, which are the most important scavengers of ozone. AsA is associated with proper seed production and, thus, woody plant reproduction. Similarly, an important function of AsA is described under drought, salinity, temperature, light stress, and biotic stress. This report emphasizes the involvement of AsA in the ecological advantages, such as nutrition recycling due to leaf senescence, of trees and shrubs compared to nonwoody plants.

Dataset Information

A possible approach for gel-based proteomic studies in recalcitrant woody plants.

Publications

A possible approach for gel-based proteomic studies in recalcitrant woody plants.

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