title: Protein extraction from Ca-alginate encapsulated plant material for comparative proteomic analysis.
abstract: The extensive use of encapsulation material in biotechnology drove the need to develop analytical techniques for this type of material. This study focuses on the specific problems of protein extraction from Ca-alginate encapsulated plant material. Proteomics is one of the fast-developing analysis categories, specifically for stress resistance and developmental changes in plant material. Sample preparation is a critical step in a two-dimensional gel electrophoresis proteome approach and is essential for good results. The aim was to avoid preliminary manipulations and get good quality material for comparative proteome analysis technique 2DE. The phenol extraction method and the complex method with preliminary TCA precipitation, SDS buffer and phenol phase were compared with respect to the efficiency and quality of the resulting 2DE gel. The most appropriate method turned out to be the TCA/phenol method with the phenol fractioning technique adapted to the gentian cell suspension. It resulted in a high protein concentration and good quality sample that could be analyzed using the standard separation procedures of 2DE and spectrometric identification with high efficiency. The work presented here confirms the possibility of obtaining a sufficient protein sample for effective proteomic analysis from a small number of capsules.
authors.affiliation: Laboratory of Plant Biotechnology, Department of Experimental Plant Biology, Polish Academy of Sciences Botanical Garden Center of Biodiversity Conservation in Powsin, 2 Prawdziwka St., 02-973, Warsaw, Poland. Electronic address: l.domzalska@obpan.pl.
authors.forename: Lucyna
authors.initials: L
authors.lastname: Domżalska
authors.affiliation: Laboratory of Plant Biotechnology, Department of Experimental Plant Biology, Polish Academy of Sciences Botanical Garden Center of Biodiversity Conservation in Powsin, 2 Prawdziwka St., 02-973, Warsaw, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Mikuła
authors.affiliation: Laboratory of Plant Biotechnology, Department of Experimental Plant Biology, Polish Academy of Sciences Botanical Garden Center of Biodiversity Conservation in Powsin, 2 Prawdziwka St., 02-973, Warsaw, Poland.
authors.forename: Jan J
authors.initials: JJ
authors.lastname: Rybczyński
mesh_terms: D000464:Alginates / Q000737:chemistry*; D030012:Gentiana / Q000737:chemistry* / Q000378:metabolism; D020723:Glucuronic Acid / Q000737:chemistry; D006603:Hexuronic Acids / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*
chemical_list: D000464:Alginates; D006603:Hexuronic Acids; D010940:Plant Proteins; D020723:Glucuronic Acid
keywords: Alginate beads; Extraction method; Gentiana cruciata; Suspension culture; Two-dimensional gel electrophoresis