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Evaluation of sample preparation protocols for proteomic analysis of sunflower leaves |
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| Authors: | Marcelo Anselmo Oseas da Silva,Jerusa Simone Garcia,Marcos Nogueira Eberlin,Marco Auré lio Zezzi Arruda |
| Affiliation: | a Group of Spectrometry, Sample Preparation and Mechanization (GEPAM), Institute of Chemistry, PO Box 6154, University of Campinas, UNICAMP, 13084-862, Campinas, SP, Brazil b National Institute of Science and Technology for Bioanalytics, Institute of Chemistry, University of Campinas, UNICAMP, PO Box 6154, 13083-970, Campinas, SP, Brazil c ThoMSon Mass Spectrometry Laboratory, Institute of Chemistry, PO Box 6154, University of Campinas, UNICAMP, 13084-862, Campinas, SP, Brazil d Department of Pharmacology, Faculty of Medical Sciences, PO Box 6111, University of Campinas, UNICAMP, 13083-887, Campinas, SP, Brazil e Department of Organic Chemistry, Institute of Chemistry, PO Box 6154, University of Campinas, UNICAMP, 13084-862, Campinas, SP, Brazil |
| Abstract: | Six protocols for extraction of proteins from sunflower (Helianthus annuus L.) leaves were evaluated for their abilities in both removing interferents and attaining the best resolution in two-dimensional gel electrophoresis. “Classical” phenol extraction followed by precipitation with ammonium acetate in methanol displayed the most efficient protocol, which allowed the detection of 244 protein spots with ca. 485 μg of protein in gel electrophoresis. Tandem mass spectrometry was performed to identify proteins in 61 spots, and cross species identification was used for this task. Proteins from twenty two spots were identified, and 12 of these proteins are up to now not included into the ExPASy sunflower protein databank. |
| Keywords: | 2D electrophoresis Sunflower Sample preparation MALDI-Q-TOF MS Cross species identification |
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