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DEVELOPMENT AND HORMONE ACTION

Proteomic Analysis of Somatic Embryogenesis in Medicago truncatula. Explant Cultures Grown under 6-Benzylaminopurine and 1-Naphthaleneacetic Acid Treatments¹

Australian Research Council Centre of Excellence for Integrative Legume Research, Genomic Interactions Group, Research School of Biological Sciences, Australian National University, Canberra City, Australian Capital Territory, 2601, Australia (N.I., M.N., B.G.R.); and University of Newcastle and Australian Research Council Centre of Excellence for Integrative Legume Research, School of Environmental and Life Sciences, University Drive, Callaghan, New South Wales, 2308, Australia (D.D., K.E.N., R.J.R.)

The Medicago truncatula line 2HA has a 500-fold greater capacity to regenerate plants in culture by somatic embryogenesis than wild-type Jemalong. We have compared proteomes of tissue cultures from leaf explants of these two lines. Both 2HA and Jemalong explants were grown on media containing the auxin 1-naphthaleneacetic acid and the cytokinin 6-benzylaminopurine. Proteins were extracted from the cultures at different time points (2, 5, and 8 weeks), separated by two-dimensional gel electrophoresis, and detected by silver staining. More than 2,000 proteins could be reproducibly resolved and detected on each gel. Statistical analysis showed that 54 protein spots were significantly (P < 0.05) changed in expression (accumulation) during the 8 weeks of culture, and most of these spots were extracted from colloidal Coomassie-stained two-dimensional gel electrophoresis gels and were subjected to matrix-assisted laser desorption ionization time-of-flight mass spectrometry or liquid chromatography-tandem mass spectrometry analysis. Using a publicly available expressed sequence tag database and the Mascot search engine, we were able to identify 16 differentially expressed proteins. More than 60% of the differentially expressed protein spots had very different patterns of gene expression between 2HA and Jemalong during the 8 weeks of culture.

Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.104.055277.

^* Corresponding author; e-mail rolfe{at}rsbs.anu.edu.au; fax 61–2–6125–0754.

Received October 19, 2004; returned for revision January 6, 2005; accepted January 9, 2005.

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