Abstract
Separation of plant proteins by means of electrophoretic techniques is quite challenging since different compounds typical for plant cells can interfere and/or reduce the effectiveness of the protein isolation. This is particularly problematic for two-dimensional electrophoresis (2-DE). Therefore, it is important to optimize protein extraction and to establish a robust protocol for 2-DE and downstream processing, primarily mass spectrometry (MS) analysis. Here we give a detailed protocol for protein extraction using phenol method, 2-DE, and MALDI-MS analysis.
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References
Oliveira BM, Coorssen JR, Martins-de-Souza D (2014) 2DE: the phoenix of proteomics. J Proteome 104:140–150. https://doi.org/10.1016/j.jprot.2014.03.035
Isaacson T, Damasceno CM, Saravanan RS et al (2006) Sample extraction techniques for enhanced proteomic analysis of plant tissues. Nat Protoc 1:769–774. https://doi.org/10.1038/nprot.2006.102
Jellouli N, Salem AB, Ghorbel A, Jouira HB (2010) Evaluation of protein extraction methods for Vitis vinifera leaf and root proteome analysis by two-dimensional electrophoresis. J Integr Plant Biol 52:933–940. https://doi.org/10.1111/j.1744-7909.2010.00973.x
Champagne A, Boutry M (2013) Proteomics of nonmodel plant species. Proteomics 13:663–673. https://doi.org/10.1002/pmic.201200312
Carpentier SC, Panis B, Vertommen A et al (2008) Proteome analysis of non-model plants: a challenging but powerful approach. Mass Spectrom Rev 27:354–377. https://doi.org/10.1002/mas.20170
Pavoković D, Križnik B, Krsnik-Rasol M (2012) Evaluation of protein extraction methods for proteomic analysis of non-model recalcitrant plant tissues. Croat Chem Acta 85:177–183. https://doi.org/10.5562/cca1804
Rogić T, Horvatić A, Tkalec M et al (2015) Proteomic analysis of Mammillaria gracilis Pfeiff. In vitro-grown cultures exposed to iso-osmotic NaCl and mannitol. Plant Cell Tissue Org 122:127–146. https://doi.org/10.1007/s11240-015-0756-9
Gundry RL, White MY, Murray CI, Kane LA, Fu Q, Stanley BA, Van Eyk JE (2009) Preparation of proteins and peptides for mass spectrometry analysis in a bottom-up proteomics workflow. Curr Protoc Mol Biol 88:VI:10.25.1–VI10.25.23. https://doi.org/10.1002/0471142727.mb1025s88
Cindrić M, Kraljević PS, Horvatić A, Dodig I (2010) Mass spectrometry-based protein identification. Patent from PCT Int. Appl. WO 2011089453
Butorac A, Dodig I, Bačun-Družina V et al (2013) The effect of starvation stress on Lactobacillus brevis L62 protein profile determined by de novo sequencing in positive and negative mass spectrometry ion mode. Rapid Commun Mass Spectrom 27:1045–1053. https://doi.org/10.1002/rcm.6528
Faurobert M, Pelpoir E, Chaïb J (2007) Phenol extraction of proteins for proteomic studies of recalcitrant plant tissues. In: Thiellement H, Zivy M, Damerval C et al (eds) Plant proteomics: methods and protocols. Humana Press, Totowa, NJ, pp 9–14. https://doi.org/10.1385/1-59745-227-0:9
Butorac A, Mekić MS, Hozić A et al (2016) Benefits of selective peptide derivatization with sulfonating reagent at acidic pH for facile matrix-assisted laser desorption/ionization de novo sequencing. Rapid Commun Mass Spectrom 30:1687–1694. https://doi.org/10.1002/rcm.7594
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Peharec Štefanić, P., Cindrić, M., Balen, B. (2018). Proteomic Analysis of Non-model Plant Tissues Using Phenol Extraction, Two-Dimensional Electrophoresis, and MALDI Mass Spectrometry. In: Loyola-Vargas, V., Ochoa-Alejo, N. (eds) Plant Cell Culture Protocols. Methods in Molecular Biology, vol 1815. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8594-4_25
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DOI: https://doi.org/10.1007/978-1-4939-8594-4_25
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