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United States Department of Agriculture

Agricultural Research Service

Title: Comparing Techniques for Comparative Proteomics: Two-Dimensional Gel Electrophoresis and Two-Dimensional Liquid Chromatography Separation

Authors
item Gunther, Nereus
item Hoan-Jen, Pang - VISITING SCI FROM RUTGERS
item Nunez, Alberto
item Uhlich, Gaylen

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: April 15, 2006
Publication Date: July 30, 2006
Citation: Gunther, N.W., Hoan-Jen, P., Nunez, A., Uhlich, G.A. 2006. Comparing techniques for comparative proteomics: two-dimensional gel electrophoresis and two-dimensional liquid chromatography separation. Society of Industrial Microbiology. {Abstract}. P75.

Technical Abstract: The accepted method for comparing bacterial proteomes has traditionally been two-dimensional (2D) gel electrophoresis. However, in recent years, new procedures for protein separation have been introduced. One of these new procedures utilizes column-based liquid chromatography separation. The techniques by which these two methods separate proteins differ significantly; however, it is currently unclear to what degree the sets of proteins identified by these different methods will diverge. To answer this question we compared the proteomes of Escherichia coli O157:H7 strain EDL933 against a variant containing a point mutation in the promoter of the csgD transcriptional regulator, using both 2D gel electrophoresis and the column-based system. Whole protein samples were prepared from the wild type and mutant and then split in half, with one half analyzed by 2D gels and the other with the columns. Differentially regulated proteins were observed in each system and identified by MALD/I-Tof/Tof analysis. The sensitivities of Coomassie blue-stained gels and the column-based system were similar, with each able to visualize and compare roughly 300 proteins. Likewise, both of the separation systems identified similar numbers of differentially regulated proteins. However, a lack of complete redundancy between the sets of proteins identified suggests that these two methods are complimentary and not strictly corroborative.

Last Modified: 12/21/2014
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