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Molecular & Cellular Proteomics 4:1812-1825, 2005.
© 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

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A Two-dimensional Electrophoresis Proteomic Reference Map and Systematic Identification of 1367 Proteins from a Cell Suspension Culture of the Model Legume Medicago truncatula^*,S

Zhentian Lei, Aaron M. Elmer, Bonnie S. Watson, Richard A. Dixon, Pedro J. Mendes and Lloyd W. Sumner^,¶

From the Plant Biology Division, The Samuel Roberts Noble Foundation, Ardmore, Oklahoma 73402 and the Virginia Bioinformatics Institute, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061

The proteome of a Medicago truncatula cell suspension culture was analyzed using two-dimensional electrophoresis and nanoscale HPLC coupled to a tandem Q-TOF mass spectrometer (QSTAR Pulsar i) to yield an extensive protein reference map. Coomassie Brilliant Blue R-250 was used to visualize more than 1661 proteins, which were excised, subjected to in-gel trypsin digestion, and analyzed using nanoscale HPLC/MS/MS. The resulting spectral data were queried against a custom legume protein database using the MASCOT search engine. A total of 1367 of the 1661 proteins were identified with high rigor, yielding an identification success rate of 83% and 907 unique protein accession numbers. Functional annotation of the M. truncatula suspension cell proteins revealed a complete tricarboxylic acid cycle, a nearly complete glycolytic pathway, a significant portion of the ubiquitin pathway with the associated proteolytic and regulatory complexes, and many enzymes involved in secondary metabolism such as flavonoid/isoflavonoid, chalcone, and lignin biosynthesis. Proteins were also identified from most other functional classes including primary metabolism, energy production, disease/defense, protein destination/storage, protein synthesis, transcription, cell growth/division, and signal transduction. This work represents the most extensive proteomic description of M. truncatula suspension cells to date and provides a reference map for future comparative proteomic and functional genomic studies of the response of these cells to biotic and abiotic stress.

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