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United States Department of Agriculture

Agricultural Research Service

Research Project: MODIFICATION OF SOYBEAN SEED COMPOSITION FOR FOOD, FEED, AND OTHER INDUSTRIAL USES

Location: Plant Genetics Research

Title: A Simple Procedure for Depletion of Storage Proteins From Soybean (Glycine max) Seeds to Aid Proteome Analysis

Authors
item Krishnan, Hari
item Oehrle, Nathan
item Natarajan, Savithiry

Submitted to: Soybean Biotechnology Meeting
Publication Type: Abstract Only
Publication Acceptance Date: April 21, 2009
Publication Date: N/A

Technical Abstract: Two-dimensional electrophoretic analysis of plant proteomes containing thousands of proteins, has limited dynamic resolution because only abundant proteins can be detected. Proteomic assessment of the low abundance proteins within seeds is difficult when 60 – 80% is storage proteins. Resolution can be improved through sample fractionation using separation techniques based upon different physiological or biochemical principles. We have developed a fast and simple fractionation technique using 10 mM Ca2+ to precipitate soybean (Glycine max) seed storage globulins, glycinin and ß-conglycinin. This method removes 87 ± 4% of the highly abundant seed proteins from the extract, allowing for 541 previously inconspicuous proteins present in soybean seed to be more detectable (volume increase of more than 50%) using fluorescent detection. Of those 541 enhanced spots, 197 increased more than 2.5-fold when visualized with Coomassie. The majority of those spots were isolated and identified using peptide mass fingerprinting. Fractionation also provided detection of 63 new phosphorylated protein spots and enhanced the visibility of 15 phosphorylated protein spots, using two-dimensional electrophoretic separation and an in-gel phosphoprotein stain. Initial experiments demonstrate possible application of this methodology towards other legumes also containing high amounts of globulin storage proteins.

Last Modified: 10/1/2014
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