Biomod/2013/OSU/protocol/folding reaction

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<img src="http://openwetware.org/images/1/15/Ohiomod2013.jpg" height="60%" width="60%" style="margin-left:-40px;"> <div id="navbar" style="margin-left:auto; margin-right:auto;text-align:center;width:650px;height:auto;" > <ul id="homebutton" class="navigation-bar"> <li><a href="http://openwetware.org/wiki/Biomod/2013/OSU">Home</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2013/OSU/about">About</a> <ul> <li><a href="http://openwetware.org/wiki/Biomod/2013/OSU/abstract">Abstract</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2013/OSU/goals">Goals</a></li>

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                       <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/folding_reaction">

<img src="http://openwetware.org/images/9/9e/Protocol_01.gif" width="129" height="125" alt=""></a></td> <td rowspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/gel_prep"> <img src="http://openwetware.org/images/4/48/Protocol_02.gif" width="99" height="123" border="0" alt=""></a></td> <td align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/negative_staining"> <img src="http://openwetware.org/images/e/ed/Protocol_03.gif" width="116" height="122" border="0" alt=""></a></td> <td rowspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/custom_tem_grids"> <img src="http://openwetware.org/images/a/ae/Protocol_04.gif" width="93" height="123" border="0" alt=""></a></td> <td colspan="2" rowspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/drugloading"> <img src="http://openwetware.org/images/e/e9/Protocol_05.gif" width="132" height="123" border="0" alt=""></a></td> <td colspan="2" rowspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/TIRF"> <img src="http://openwetware.org/images/f/fd/Protocol_06.gif" width="137" height="123" border="0" alt=""></a></td> <td rowspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/MTT"> <img src="http://openwetware.org/images/2/20/Protocol_07.gif" width="94" height="123" border="0" alt=""></a></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="1" height="122" alt=""></td> </tr> <tr> <td rowspan="3"> <img src="http://openwetware.org/images/0/0b/Protocol_08.gif" width="116" height="3" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="1" height="1" alt=""></td> </tr> <tr> <td rowspan="2"> <img src="http://openwetware.org/images/5/59/Protocol_09.gif" width="99" height="2" alt=""></td> <td colspan="2" rowspan="2"> <img src="http://openwetware.org/images/e/e4/Protocol_10.gif" width="211" height="2" alt=""></td> <td colspan="2" align="left" valign="top"> <a href="http://openwetware.org/wiki/Biomod/2013/OSU/protocol/cellanalysis"> <img src="http://openwetware.org/images/3/3c/Protocol_11.gif" width="108" height="1" border="0" alt=""></a></td> <td colspan="2" rowspan="2"> <img src="http://openwetware.org/images/8/83/Protocol_12.gif" width="137" height="2" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="1" height="1" alt=""></td> </tr> <tr> <td colspan="2"> <img src="http://openwetware.org/images/7/77/Protocol_13.gif" width="108" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="1" height="1" alt=""></td> </tr> <tr> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="129" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="99" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="116" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="93" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="118" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="14" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="94" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="43" height="1" alt=""></td> <td> <img src="http://openwetware.org/images/5/52/Spacer.gif" width="94" height="1" alt=""></td> <td></td> </tr> </table> </center>

<div id="protocol-content" style="width:65%;margin-left:auto;margin-right:auto;"> <h1>Folding Reaction</h1> <p>A folding reaction is a mixture containing scaffold DNA, staple DNA, water, a buffer mastermix, and magnesium ions. The folding involves subjecting the sample mixture to thermal denaturation followed by renaturing. We will usually have the folding reactions occur in PCR strip tubes of either 50-100 μL aliquots.</p>

<h2>Pre-stocks</h2> <p>A folding reaction is a mixture containing scaffold DNA, staple DNA, water, a buffer mastermix, and magnesium ions. The folding involves subjecting the sample mixture to thermal denaturation followed by renaturing. We will usually have the folding reactions occur in PCR strip tubes of either 50-100 μL aliquots.</p>

<table> <tr> <td><b>Index</b></td> <td><b>Plate</b></td> <td><b>From Well</b></td> <td><b>To Well</b></td> <td><b>Name</b></td> <td><b>No. of Oligos</b></td> <td><b>Conc. (um)</b></td> <td><b>Element</b></td> </tr> <tr> <td>p01</td> <td>p085</td> <td>A1</td> <td>H12</td> <td>CoreP085</td> <td>96</td> <td>1.04</td> <td>core 1</td> </tr> <tr> <td>p02</td> <td>p085</td> <td>A1</td> <td>H12</td> <td>CoreP086</td> <td>41</td> <td>2.44</td> <td>core 2</td> </tr> <tr> <td>p03</td> <td>p086</td> <td>E1</td> <td>F1</td> <td>Neighbors</td> <td>13</td> <td>7.69</td> <td>Neighbors</td> </tr> <tr> <td>p04</td> <td>p086</td> <td>G1</td> <td>H1</td> <td>Polymers</td> <td>13</td> <td>7.69</td> <td>Replaced by polymers</td> </tr> </table>

<h2>Working Stocks</h2> <p>After pipetting the pre-stocks (hint: use multi-channel pipette) we will now take aliquots from each pre-stock to form the working stock. <a href="http://openwetware.org/images/7/79/Pre_stocks_5-24_origamiv3.xls">Here</a> is an example sheet used.</p>

<table> <tr> <td><b>Pre-stock Source</b></td> <td><b>#</b></td> <td><b></b></td> <td><b>[uL]</b></td> <td><b></b></td> <td><b>[uL]</b></td> </tr> <tr> <td>CoreP085</td> <td>96</td> <td>1</td> <td>96</td> <td>1</td> <td>96</td> </tr> <tr> <td>CoreP086</td> <td>41</td> <td>1</td> <td>41</td> <td>1</td> <td>41</td> </tr> <tr> <td>Neighbors</td> <td>13</td> <td>1</td> <td>13</td> <td>0</td> <td>0</td> </tr> <tr> <td>Polymers</td> <td>13</td> <td>1</td> <td>13</td> <td>0</td> <td>0</td> </tr> <tr> <td>ddH20</td> <td></td> <td></td> <td>37</td> <td></td> <td>63</td> </tr> <tr> <td><b>Sum Volume</b></td> <td></td> <td></td> <td>200</td> <td></td> <td>200</td> </tr> <tr> <td>Concentration</td> <td></td> <td></td> <td>500 nM</td> <td></td> <td>500 nM</td> </tr> </table>

<h2>Prepare Folding Reaction</h2> <p>Now that we have our working stock, we must decide whether to use 10 nM or 20 nM of our scaffold concentration, and whether hte final volume will be 50 μL or 100 μL. By default the FOBXM buffer solution and the MgCl2 magnesium screen will have already been prepared.</p>

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