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Two Dimensional Electrophoresis Analysis of the Proteins Responsive to Gravity in Caenorhabditis elegans.  A. Higashibata1, M. Imamizo-Sato2, M. Seki2 and N. Ishioka1.  1Department of Space Biology and Microgravity Sciences, Institute of Space and Astronautical Science, Japan Aerospace Exploration Agency, 2Space Development Division, Advanced Engineering Services Co., Ltd, Tsukuba, Ibaraki, JAPAN.

   In May of this year, we participated in ICE-First that was the international cooperative flight experiment, and we acquired the flight sample of C. elegans.  We focus on the investigation of the effects of gravity on protein expression in C. elegans.  The whole lysate was prepared from three experimental groups; ground control, clinorotation and flight.  The proteins were extracted from each group using CHAPS buffer (8M Urea, 2% CHAPS, 50mM DTT, 0.2% Bio-Lytes 3-10, 0.001% BPB).  2D electrophoresis was performed using Bio-Rad System.  The lysate containing 500 ?g of total proteins was loaded onto an immobilized pH3-10 linear gradient strip, and focused to 20,000 Vh.  Then the strip applied to a 10-20% gradient gel.  The total proteins were detected using SYPRO Ruby.  The phosphoproteins and the glycoproteins were detected by Pro-Q Diamond and Pro-Q Emerald staining, respectively.  The images were analyzed using PD Quest software.  On the gel of SYPRO Ruby staining, several spots indicating different expression level were detected in the area of pI6-8 and Mw 20,000-50,000.  These certain spots are isolated from the gel and extracted to identify by mass spectrometry.