Biomod/2014/Hokudai/MATERIAL&METHOD
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<nobr> <p class="example111 example2 example3 example4 example5"> <a href="PROJECT">PROJECT</a> <a href="DESIGN">DESIGN</a> <a href="MATERIAL&METHOD">MATERIAL&METHOD</a> <a href="RESULTS">RESULTS</a> <a href="FUTURE">FUTURE</a> <a href="TEAM">TEAM</a> <a href="SPONSORS">SPONSORS</a> </nobr> </p>
<p class="example7">Term</p> <p class="example8">Microtubule</p> <p class="example6">Microtubules are filamentous and cylindrical biopolymers of the heterodimer of tubulin. They are present in virtually all eukaryotic cells as a major component of an interconnected network of filaments known as the cytoskeleton. They perform various functions not only in cell motility, cell division, organization and orientation but also in transporting organelles as rail for motor proteins.
<p class="example8">Kinesin</p> <p class="example6">Kinesin is a motor protein found in eukaryotic cells. It moves along microtubule filaments fueled by the hydrolysis of ATP. It helps transport cellular cargoes and support several cellular functions including mitosis, meiosis and cytokinesis.</p> <p class="example8">Sarcomere</p> <p class="example6">Sarcomere is the contractile unit of muscle. It is comprised of actin filaments and bipolar myosin filaments (multimeric myosins) which are organized into high oriented structure. Sliding motion between actin filaments and myosin filaments causes shortening all the sarcomere and subsequently the entire muscle. </p>
<p class="example7">Process</p>
<p class="example6"><br>① We prepared flow cells by placing a cover glass (18 × 18 mm2; MATSUNAMI) on an another slide glass (26 × 76 mm2) with a pair of double sided tapes as spacers. Size of flow cell: 1.5 × 18 × 0.1 mm3 (~3 µL) (W × L × H) in dimension.</br>
<br>② GTP-buffer and tubulin solution were mixed at the ratio of 1 : 4 (Final tubulin concentration: 112.5 µM).</br>
<br>③ Tubulin mixture was incubated at 4 °C for 10 minutes to depolymerize it completely. It was confirmed that polymerization of tubulin had not been started yet by using a polarizing microscopes.</br>
<br>④ The backside of the area of interest in the flow cell was fixed with a black tape where polymerization of tubulin would be initiated by IR irradiation. </br>
<br>⑤ A part of the flow cell where black tape was fixed was warmed for 15 minutes by using IR irradiation (IRS-001C; IR SYSTEM Co., Ltd., 5V / 1.6A).</br>
<br>⑥ Microtubules were observed by using fluorescence* microscope and polarizing microscope.</br>
<br>*That time we used TAMRA labeled tubulin.</br>
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