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Control of Power Generation in Actin-Myosin Gels Yamamoto Sho
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Way of Control over Power Generation in Actin-Myosin gels 3、 exchange used dialysis membrane 4、 Photolysis of caged 1、 Temperature adjustment in the sample generation 2、 ATP regeneration system used PC, CPK
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1、 Temperature adjustment in the sample generation ・ All the preparations and implements were kept to be cold. ( about at 0 ℃) ・ When observing,I put the samples on the THERMO PLATE which was set at 3 ℃
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Result At room temperature(24 ℃ ) On THERMO PLATE
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Problem ・ I could not control the exact temperature of the “actin-myosin gels”. ・ The probe particles swayed from one side to another.(Probably because of the watere current in the THERMO PLATE.) Solution ・ To cover a box to keep the inner temperature cold. ・ Not to use oil immersion lens. ( The oil conduct heat of the objective lens to the samples.)
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exchange used dialysis membrane3. Jan Scrimgeour,*ab Jae Kyu Cho,c Victor Breedveldc and Jennifer Curtisab,Soft Matter,2011
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Problem Prepolymer solution did not be polymerized. Polyethylene glycol diacrylate has MEHQ as inhibitor. Solution Distillation under reduced pressure Boiling point of MEHQ : 243 ℃ 760mmHg 126 ℃ 11mmHg 5.Cooling water 9.Vacuum pump
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In the cell, force generation on actin-myosin network is controlled by calcium ion concentration. After released by endoplasmic reticulum, calcium is combined with troponin, and tropomyosin is caused a structural change, and finally actin is made activated. Function ofin Muscular Tissue
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Method ・ F-actin and Tn-Tm complex are mixed to final concentrations of 1mg/ml each in the presence of 0.1M KCl, 0.1mM ATP, 10mM-TrisHCl(pH=8.0) and the mixture are heated at 45 ℃ for 15 min and then cool at a slow rate (about 0.3 deg/min) to room temperature. To confirm F-actin is crosslinked with TN-TM, I should observe superprecipitation in the presence of Ca ion. Hajime Honda7 and Sho Asakura,1988
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(a) and (c) Video-recorded images of fluorescent actin filaments and (b) and (d) changes in position of the actin filaments brought about by their movement in 0.6 s at (a) and (b) pCa = 3, (c) and (d) pCa = 5.8 at 30 ℃. Hajime Honda7 and Sho Asakura,1988
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The average verocities at pCa=3 of regulated actin(continuous line with filled circle) and unregulated actin(broken line with open triangles).The broken line with open circle shows the temperature-dependence of regulated actin-activated myosin ATPase activity.
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