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Poster De Conférence Année : 2023

Evaluation of the efficiency of the SPEED protein extraction protocol on plant samplesEvaluation of the efficiency of the SPEED protein extraction protocol on plant samples

Résumé

In bottom-up proteomics, sample preparation is a critical step that strongly influences the quality of the results. In particular, protein extraction should enable efficient protein digestion for subsequent mass spectrometric analysis. Recently, Doellinger et al. [1] reported a detergent-free method called Sample Preparation by Easy Extraction and Digestion (SPEED), which consists of three mandatory steps: acidification, neutralization, and digestion. SPEED is an attractive method that holds the potential to dramatically simplify and standardize sample preparation while improving the depth of proteome coverage, especially for challenging samples. In addition, its authors report it as universal because it uses pure trifluoroacetic acid, which is supposed to extract protein from any sample by complete dissolution. However, this method has not been tested on plant samples. Yet, plants have specific compounds (e.g., polyphenols, chlorophyll, or major proteins) that can interfere with sample preparation. Here, we aim to evaluate if the SPEED method is suitable for preparing plant protein extracts. To this end, we sampled leaves, roots, and seeds from Arabidopsis Col-0 plants. We ground each tissue in liquid nitrogen and divided the resulting powders into nine aliquots that we subsequently used to extract proteins by three different protocols in three replicates: the SPEED method, the TCA-acetone-based, and the phenol-based extraction methods. We routinely use the latter two protocols to extract proteins from leaves and seeds, respectively. Mass spectrometry analysis for the whole experimental design is currently in progress. Preliminary results obtained on leaf samples indicate that the SPEED method provides good extraction yields and mass spectrometry signals. [1] Doellinger et al. (2020) Sample Preparation by Easy Extraction and Digestion (SPEED) - A Universal, Rapid, and Detergent-free Protocol for Proteomics Based on Acid Extraction. Mol Cell Proteom.(1):209-222. doi: 10.1074/mcp.TIR119.001616.
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Dates et versions

hal-04302587 , version 1 (23-11-2023)

Identifiants

  • HAL Id : hal-04302587 , version 1

Citer

Thierry Balliau, Shabnam Huseynova, Marlène Davanture, Melisande Blein-Nicolas. Evaluation of the efficiency of the SPEED protein extraction protocol on plant samplesEvaluation of the efficiency of the SPEED protein extraction protocol on plant samples. ProteoAix, Jun 2023, Aix-en-Provence, France. ⟨hal-04302587⟩
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