Archived Protocol

<!DOCTYPE HTML PUBLIC "-//W3C//DTD HTML 4.0 Transitional//EN"> <html><head> <title>   Single component | molbiol.ru   </title> <meta http-equiv=keywords name=keywords content="organic, pKa, ATP, Betaine, Cresol, red, DTT, dNTP's, EDTA, EtBr, Gelatin, Glucose, Guanidine, HEPES, Imidazol, Paraformaldehyde, PEG, PMSF, Retinoic acid, Sucrose, Tris, Tricine, Triethanolamine, Urea, acids, alkalis, NaOH, KOH, ÒÕÓ, detergents, Lauroylsarcosine, SDS, solution, phenol, EtOH, ethanol, DNA, RNA, protein, gene, vector, genome, bacteria, virus, cell, cloning, experiment, methods, tip, trick, expression, method, recipies, calculations"> <meta http-equiv=description name=description content="Composition of molecular biology solutions. Single component solutions. Practical Molecular Biology."> <meta name="Document-State" content="Static"> <meta name="Robots" content="ALL"><meta name="Robots" content="ALL"> <LINK rel="stylesheet" type="text/css" href="/style.css"> </head> <body background="/izo/fon.gif" topmargin="0" leftmargin="0" marginwidth="0" marginheight="0"> <a name="start"></a> <table width="800" border="0" cellspacing="0" cellpadding="0"> <tr><td colspan="2" width="610" valign="top">  <a href=http://molbiol.ru/eng/index.html><img src=http://molbiol.ru/izo/logo.gif width="98" height="41" alt="molbiol.ru - main page." border="0" align=left></a><img src=http://molbiol.ru/izo/1.gif width="13" height="13" border="0"><br><font color="Gray" size="-1"> > <a href=http://molbiol.ru/eng/protocol/index.html>Protocols</a> >  <a href=http://molbiol.ru/eng/protocol/index.html>Solutions</a> > <b>Single component.</font></b> <hr align="left" size="2" color="Gray" noshade> <p class="h1"> Single component solutions. </p></td><td rowspan="3" width="190" valign="top"> <div class="enav"><br> <FORM METHOD="GET" ACTION="/cgi/gliws.pl/frm/search"> <INPUT TYPE="text" NAME="query" SIZE=15><br> <INPUT TYPE="submit" NAME="Search" VALUE="Find"><br> <hr align="left" width="125" size="2" color="Gray" noshade> <a href=http://molbiol.ru/solution/03_02.html><img src=http://molbiol.ru/izo/rus.gif width="32" height="21" alt="Russian version" border="0"></a>  <a href=http://molbiol.ru/ger/protocol/01_03.html><img src=http://molbiol.ru/izo/ger.gif width="32" height="21" alt="Deutsche Version" border="0"></a>  <a href=http://molbiol.ru/ukr/protocol/01_03.html><img src=http://molbiol.ru/izo/ukr.gif width="32" height="21" alt="Óêðà¿íñüêèé âàð³àíò" border="0"></a>  <br><hr align="left" width="125" size="2" color="Gray" noshade> <a href=http://molbiol.ru/eng/index.html>Main page.</a><br> <a href=http://molbiol.ru/eng/project.html>Project.</a><br> <a href=http://molbiol.ru/eng/protocol/><b>Protocols.</b></a><br>    <a href=http://molbiol.ru/eng/protocol/index.html><font size="-1"><b>§ Solutions.</b></font></a><br> <a href=http://molbiol.ru/eng/scripts/>Calculations.</a><br> <a href=http://molbiol.ru/cgi-bin/biomail/users.pl>BioMail.</a><br> <a href=http://molbiol.ru/eng/journals/>Literature.</a><br> <a href=http://molbiol.ru/eng/workdb/>Labor exchange.</a><br> </FORM>  <table width="100" border="1" cellpadding="3" bordercolorlight="#FFFFFF"><tr align="center" valign="middle" bgcolor="#FFFFFF"><td><a href=http://molbiol.ru/review/index.html#_2a>Ñòðóêòóðíàÿ îðãàíèçàöèÿ áåëêà<br>Å.Ì.Ïîïîâ</td></tr></table> </div> <img src=http://counter.rambler.ru/top100.cnt?164472 alt="Rambler's Top100" width=1 height=1 border=0><img height=1 width=1 src=http://top.list.ru/counter?id=339529><img src=http://cnt.one.ru/cgi-bin/cnt.cgi?id=145057 width=1 height=1 alt="" border=0> <script language="javascript"></script><noscript> <img src=http://u586.76.spylog.com/cnt?cid=58676&p=1 alt='SpyLOG' border='0' width=1 height=1 > </noscript>  </td></tr> <tr><td width="310" bgcolor="eeeeee" valign="top"> <script language="JavaScript" src=http://molbiol.ru/eng/scripts/sol_a.js type="text/javascript"></script> <div class="inav"> <A href=http://molbiol.ru/eng/protocol/01_03.html#a1>Organic substances.</A><br> <div class="ls"><A href=http://molbiol.ru/eng/protocol/01_03.html#a2>pK<sub>a</sub> and temperature dependence of pH for common buffers.</A><br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a3>ATP</A> 0.1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a4>Betaine</A> 5M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a5>Cresol red (Na)</A> 50mM<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a6>DTT</A> 1M, 2.2M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a7>dNTP’s</A> 100mM<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a8>EDTA</A> 0.5M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a9>EtBr</A> 10mg/ml<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a10>Gelatin</A> 2%<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a11>Glucose</A> 1; 1.5; 2M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a12>Guanidine HCl</A> 1-8M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a13>HEPES</A> 1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a14>Imidazol</A> 2M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a15>Paraformaldehyde</A> 37%<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a16>PEG</A> 40%<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a17>PMSF</A> 100mM<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a18>Retinoic acid</A> 10mM<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a18a>Sucrose</A> 1; 2; 2.5M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a19>Tris Cl</A> 1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a20>Temperature dependence of pH for TrisCl.</A><br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a21>Tricine</A> 1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a22>Triethanolamine</A> 1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a23>Urea</A> 1-10M<br></div> <A href=http://molbiol.ru/eng/protocol/01_03.html#a24>Acids and alkalis.</A><br> <div class="ls"><A href=http://molbiol.ru/eng/protocol/01_03.html#a24>Summary table.</A><br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a25>NaOH</A> 10M, 1M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a26>KOH</A> 5M<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a27>TCA</A> 100%<br></div> <A href=http://molbiol.ru/eng/protocol/01_03.html#a28>Detergents.</A><br> <div class="ls"><A href=http://molbiol.ru/eng/protocol/01_03.html#a29>N-Lauroylsarcosine Na</A> 10%<br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a30>SDS</A> 10%<br></div> <A href=http://molbiol.ru/eng/protocol/01_03.html#a31>Organic solvents.</A><br> <div class="ls"><A href=http://molbiol.ru/eng/protocol/01_03.html#a32>Phenol</A><br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a33>EthanolEtOH</A><br> <A href=http://molbiol.ru/eng/protocol/01_03.html#a34>Preparation of 100% EtOH.</A><br></div> <A href=http://molbiol.ru/eng/protocol/01_03.html#add><img src=http://molbiol.ru/izo/k.gif width="11" height="9" alt="" border="0"> Supplement.</A><br> </div> </TD><td valign="top"> <div class="epig">Densities of some solutions are available on the page "<a href=http://molbiol.ru/eng/protocol/01_23.html target="_blank">Densities of acids, alkali and organic substances</a>". </div> <ul type="disc"><b>See also:</b><br> <li><font size="-1"><a href=http://www.embl-heidelberg.de/JaMBW/5/4/index.html target="_blank" title="Ðàñ÷¸ò áóôåðîâ.">Buffer design</a> - online calculations of large list of buffers (Prof. R.Beynon) on the page <a href=http://www.embl-heidelberg.de/JaMBW/ target="_blank" title="Java programs for molecular biology (EMBL).">Java based Molecular Biologist's Workbench</a> EMBL.</li> <li><a href=http://www.labvelocity.com/tools/bufferCalculator.jhtml target="_blank" title="Ðàñ÷¸ò áóôåðîâ.">Buffer Calculator</a> on the site of LabVelocity. Registration is necessary (free).</li> </font></ul> <br><br><br><br><br> <ul><form name="hlp" id="hlp"><center>About the recalculation of recipes for the arbitrary volumes:<br><br><INPUT TYPE="button" VALUE="Help" onClick="helpw()"></center></form></ul> </td></tr> <tr><td width=610 colspan="2" valign="top"> </td></tr></table> <table width="610" border="0" cellspacing="0" cellpadding="0"><tr><td> <script language="JavaScript" src=http://molbiol.ru/scripts/solution.js type="text/javascript"></script> <div class="prot"><br><ul> <A name="a1"></A><p class=h2>Organic substances.</P> <p class=h3><A name="a2"></A>pK<sub>a</sub> and temperature dependence of pH for common buffers.</P> <TABLE width="550" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"><TD> <TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><TD>Buffer</TD><td>Mw</td><TD>pK<SUB>a</SUB><br>20<SUP>o</SUP>C</TD><td>Working<br>range</td><TD>delta pK<SUB>a</SUB><br>per 10<SUP>o</SUP>C</TD></TR> <TR bgcolor="ffffff"><TD><b>MES</b><br><font size="-1">2-(<i>N</i>-morpholino)ethanesulfonic acid</font></TD><td>195.2</td><TD>6.15</TD><td>5.5-6.7</td><TD>-0.110</TD></TR> <TR bgcolor="ffffff"><TD><i><b>Bis-Tris</b></i><br><font size="-1"><i>bis</i>(2-hydroxyethyl)imino<i>tris</i>(hydroxymethyl)methane</font></TD><td>209.2</td><TD>6.5</TD><td>5.8-7.2</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>ADA</b><br><font size="-1"><i>N</i>-(2-acetamido)-2-imidoacetic acid</font></TD><td>190.2</td><TD>6.60</TD><td>6.0-7.2</td><TD>-0.110</TD></TR> <TR bgcolor="ffffff"><TD><b>PIPES</b><br><font size="-1">piperasine-N,N'-<i>bis</i>(2-ethanesulfonic acid)</font></TD><td>302.4</td><TD>6.80</TD><td>6.1-7.5</td><TD>-0.085</TD></TR> <TR bgcolor="ffffff"><TD><b>ACES</b><br><font size="-1">2-[(2-amino-2-oxoethyl)amino]ethanesulfonic acid</font></TD><td>182.2</td><TD>6.90</TD><td>6.1-7.5</td><TD>-0.200</TD></TR> <TR bgcolor="ffffff"><TD><b>MOPSO</b><br><font size="-1">3-(<i>N</i>-morpholino)-2-hydroxypropanesulfonic acid</font></TD><td>225.3</td><TD>6.9</TD><td>6.2-7.6</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>Imidazol - HCl</b></TD><td>68.08</td><TD>6.95</TD><td>6.2-7.8</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b><i>Bis-Tris</i> Propane</b><br><font size="-1">1,3-<i>bis</i>[<i>tris</i>(hydroxymethyl)methylamino]propane</font></TD><td>282.3</td><TD>6.8*</TD><td>6.3-9.5</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>BES</b><br><font size="-1"><i>N,N-bis</i>(2-hydroxyethyl)-2-aminoethanesulfonic acid</font></TD><td>213.2</td><TD>7.15</TD><td>6.4-7.8</td><TD>-0.160</TD></TR> <TR bgcolor="ffffff"><TD><b>MOPS</b><br><font size="-1">3-(<i>N</i>-morpholino)propanesulfonic acid</font></TD><td>209.3</td><TD>7.20</TD><td>6.5-7.9</td><TD>-0.013</TD></TR> <TR bgcolor="ffffff"><TD><b>TES</b><br><font size="-1"><i>N-tris</i>(hydroxymethyl)methyl-2-aminoethanesulfonic acid</font></TD><td>229.2</td><TD>7.50</TD><td>6.8-8.2</td><TD>-0.200</TD></TR> <TR bgcolor="ffffff"><TD><b>HEPES</b><br><font size="-1"><i>N</i>-(2-hydroxyethyl)piperazine-<i>N'</i>-(2-ethanesulfonic acid)</font></TD><td>238.3</td><TD>7.55</TD><td>6.8-8.2</td><TD>-0.140</TD></TR> <TR bgcolor="ffffff"><TD><b>DIPSO</b><br><font size="-1">3-[<i>N,N-bis</i>(2-hydroxyethyl)amino]-2-hydroxyprpanesulfonic acid</font></TD><td>243.3</td><TD>7.6</TD><td>7.0-8.2</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>TAPSO</b><br><font size="-1">3-[<i>N-tris</i>(hydroxymethyl)methylamino]-2-hydroxypropanesulfonic acid</font></TD><td>259.3</td><TD>7.6</TD><td>7.0-8.2</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>HEPPSO</b><br><font size="-1"><i>N</i>-(2-hydroxyethyl)piperazine-<i>N'</i>-(2-hydroxypropanesulfonic acid)</font></TD><td>268.3</td><TD>7.8</TD><td>7.1-8.5</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>POPSO</b><br><font size="-1">piperazine-<i>N,N'-bis</i>(2-hydroxypropanesulfonic acid)</font></TD><td>362.4</td><TD>7.8</TD><td>7.2-8.5</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>TEA</b><br><font size="-1">triethanolamine</font></TD><td>149.2</td><TD>7.8</TD><td>7.3-8.3</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>EPPS</b><br><font size="-1"><i>N</i>-(2-hydroxyethyl)piperazine-<i>N'</i>-(3-propanesulfonic acid)</font></TD><td>252.3</td><TD>8.0</TD><td>7.3-8.7</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>Tricine</b><br><font size="-1"><i>N-tris</i>(hydroxymethyl)methylglycine</font></TD><td>179.2</td><TD>8.15</TD><td>7.4-8.8</td><TD>-0.210</TD></TR> <TR bgcolor="ffffff"><TD><b>Tris (TRIZMA)</b><br><font size="-1"><i>tris</i>(hydroxymethyl)aminomethane</font></TD><td>121.1</td><TD>8.30</TD><td>7.0-9.1</td><TD>-0.310</TD></TR> <TR bgcolor="ffffff"><TD><b>Bicine</b><br><font size="-1"><i>N,N-bis</i>(2-hydroxyethyl)glycine</font></TD><td>163.2</td><TD>8.35</TD><td>7.6-9.0</td><TD>-0.180</TD></TR> <TR bgcolor="ffffff"><TD><b>TAPS</b><br><font size="-1"><i>N-tris</i>(hydroxymethyl)methyl-3-aminopropanesulfonic acid</font></TD><td>243.3</td><TD>8.4</TD><td>7.7-9.1</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>Glycylglycine</b></TD><td> </td><TD>8.40</TD><td> </td><TD>-0.280</TD></TR> <TR bgcolor="ffffff"><TD><b>AMPSO</b><br><font size="-1">3-[(1,1-dimethyl-2-hydroxyethyl)amino]-2-hydroxypropanesulfonic acid</font></TD><td>227.3</td><TD>9.0</TD><td>8.3-9.7</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>CHES</b><br><font size="-1">1-(<i>N</i>-cyclohexylamino)ethanesulfonic acid</font></TD><td>207.3</td><TD>9.3</TD><td>8.6-10.0</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>CAPSO</b><br><font size="-1">3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid</font></TD><td>237.3</td><TD>9.6</TD><td>8.9-10.3</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>AMP</b><br><font size="-1">2-amino-2-methyl-1-propanol</font></TD><td>89.1</td><TD>9.7</TD><td>9.0-10.5</td><TD> </TD></TR> <TR bgcolor="ffffff"><TD><b>CAPS</b><br><font size="-1">3-(cyclohexylamino)-1-propanesulfonic acid</font></TD><td>221.3</td><TD>10.4</TD><td>9.7-11.1</td><TD> </TD></TR> </TABLE></TD></TR> </TABLE><br>* pKa=9.0 for the second stage of dissociation. <form><A name="a3"></A><p class=h3>ATP</P> <P>C<SUB>10</SUB>H<SUB>14</SUB>N<SUB>5</SUB>O<SUB>13</SUB>P<SUB>3</SUB>Na<SUB>2</SUB> Mw=551.1 g/M; (store at -20<SUP>o</SUP>C).</P> <TABLE width="229" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"><TD> <TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>1ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>ATP</TD><TD>0.1M</TD><TD>551.1g/M</TD><TD>55.1mg</TD><TD><input type="text" size=7 onChange='clc(this,1)'></TD></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD> </TD><td> </td></TR> </TABLE></TD></TR> </TABLE> <P><img src=http://molbiol.ru/izo/trefa.gif alt='Unchecked information' width="11" height="9" alt="" border="0"> 0.1M ATP (pH 7.2): 5.71% (w/w) ATP, 84.86% (w/w) H<SUB>2</SUB>O, 9.43% (w/w) 2M Tris base.</P> <ol><li>adjust to pH7.5 by 2M NaOH (~70-80µl);</li> <li>prepare about 700µl, dilute 2000 times (it is the final dilution in the spectrofotometric cell), check the optical density:</li> <p>C[M]=A<SUB>259</SUB>/15.4x10<SUP>3</SUP>=A<SUB>259 1:2000</SUB>x0.130</p> <li>adjust the final volume.</li> </ol><input type="hidden" name="prm" value="1,2,ml,55.11,1,mg"></form> <form><A name="a4"></A><p class=h3>Betaine</P> <P>monohydrate C<SUB>5</SUB>H<SUB>11</SUB>NO<SUB>2</SUB>xH<SUB>2</SUB>O; Mw=135.2g/M; (store at 4<SUP>o</SUP>C).</P> <TABLE width="500" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>%(w/w)</TD><TD>100ml</TD><TD>200ml</TD><TD>700ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>Betaine</TD><TD>135.2g/M</TD><TD>5M</TD><TD>63.00</TD><TD>67.6g</TD><TD>135.2g</TD><TD>473.2g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD>37.00</TD><TD>39.74ml</TD><TD>79.49ml</TD><TD>278.2ml</TD><td><input type="text" size=7 onChange='clc(this,2)'></td></TR> </TABLE></TD></TR> </TABLE> <P>p~1.073g/ml</P><input type="hidden" name="prm" value="100,2,ml,67.6,2,g,39.74,2,ml"></form> <form><A name="a5"></A><p class=h3>Cresol red (Na)</P> <P>50mM (store at +4, -20<SUP>o</SUP>C):</P> <TABLE width="450" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>1ml</TD><TD>50ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>Cresol red (Na)</TD><TD>50mM</TD><TD>404.4g/M</TD><TD>20.2mg</TD><TD>1.01g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD> </TD><TD> </TD><TD> </TD></TR> </TABLE></TD></TR> </TABLE> <li>Cresol Red (Na salt) is a very convenient dye. At a concentration of about 0.2mM it is compatible with restriction digestion, PCR, sequencing. It may be used as marker for electrophoresis;</li> <li>the color of the dye is pH-dependent (red, if pH>7.5; yellow, if pH <7.0). It is possible to use Cresol Red as pH-indicator (i) for denaturation of double-stranded DNA in sequencing, (ii) for silica-purification of DNA from agarose gel.</li> <input type="hidden" name="prm" value="100,2,ml,2.022,2,g"></form> <form><A name="a6"></A><p class=h3>DTT</P> <P>C<SUB>4</SUB>H<SUB>10</SUB>O<SUB>2</SUB>S<SUB>2</SUB> Mw=154.2g/M; (store at -20<SUP>o</SUP>C).</P> <TABLE width="550" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>%(w/w)</TD><TD>1ml</TD><TD>5ml</TD><TD>10ml</TD><TD>20ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>DTT</TD><TD>154.2g/M</TD><TD>1M</TD><TD>14.55</TD><TD>0.155g</TD><TD>0.773g</TD><TD>1.55g</TD><TD>3.09g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>AcNa, pH 5.2</TD><TD> </TD><TD>10mM</TD><TD>85.45</TD><TD>905 µl</TD><TD>4.53ml</TD><TD>9.05ml</TD><TD>18ml</TD><td><input type="text" size=7 onChange='clc(this,2)'></td></TR> </TABLE></TD></TR> </TABLE> <P>p~1.06g/ml</P><input type="hidden" name="prm" value="100,2,ml,15.42,2,g,90.58,2,ml"></form> <form><TABLE width="500" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>1ml</TD><TD>5ml</TD><TD>10ml</TD><TD>20ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>DTT</TD><TD>154.2g/M</TD><TD>2.2M</TD><TD>0.339g</TD><TD>1.696g</TD><TD>3.394g</TD><TD>6.787g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>AcNa, pH 5.2</TD><TD> </TD><TD>10mM</TD><TD> </TD><TD> </TD><TD> </TD><TD> </TD><td> </td></TR> </TABLE></TD></TR> </TABLE> <li>filter sterilization;</li> <li>no DEPC treatment.</li> <input type="hidden" name="prm" value="100,2,ml,33.924,2,g"></form> <A name="a7"></A> <p class=h3>Preparation of dNTP's.</P> <P><em>The quick protocol for ~100mM stock.</em></P> <OL> <LI>dilute all four dNTP's (250mg of each) in 3.676ml of H<SUB>2</SUB>O;</LI> <LI>+ 424µl 5M NaOH;</LI> <LI>check the pH: ~0.5µl on pH-paper.</LI> </OL><br> <P><em>Accurate protocol for 100mM stocks preparation.</em></P> <OL> <LI>add the necessary quantity (see table) of H<SUB>2</SUB>O and Tris base 1M (it is possible to take the volume of salt as ~150µl);</LI> <TABLE width="456" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td> </td> <TD> </TD><TD><P>Mw</TD><TD>V<SUB>(H2O)</SUB></TD><TD>V<SUB>(Tris base 1M)</SUB></TD><TD>V<SUB>(final)</SUB></TD></TR> <TR bgcolor="ffffff"><TD>dATP</TD><TD>C<SUB>10</SUB>H<SUB>14</SUB>N<SUB>5</SUB>O<SUB>12</SUB>P<SUB>3</SUB>Na<SUB>2</SUB>x3H<SUB>2</SUB>O</TD><TD>589.2</TD><TD>3.24ml</TD><TD>850 µl</TD><TD>4.24ml</TD></TR> <TR bgcolor="ffffff"><TD>dGTP</TD><TD>C<SUB>10</SUB>H<SUB>13</SUB>N<SUB>5</SUB>O<SUB>13</SUB>P<SUB>3</SUB>Na<SUB>3</SUB>x2H<SUB>2</SUB>O</TD><TD>609.2</TD><TD>3.55ml</TD><TD>400 µl</TD><TD>4.10ml</TD></TR> <TR bgcolor="ffffff"><TD>dCTP</TD><TD>C<SUB>9</SUB>H<SUB>13</SUB>N<SUB>3</SUB>O<SUB>13</SUB>P<SUB>3</SUB>Na<SUB>3</SUB>x2H<SUB>2</SUB>O</TD><TD>569.1</TD><TD>3.44ml</TD><TD>800 µl</TD><TD>4.39ml</TD></TR> <TR bgcolor="ffffff"><TD>dTTP</TD><TD>C<SUB>10</SUB>H<SUB>14</SUB>N<SUB>2</SUB>O<SUB>14</SUB>P<SUB>3</SUB>Na<SUB>3</SUB>x2H<SUB>2</SUB>O</TD><TD>584.1</TD><TD>3.73ml</TD><TD>400 µl</TD><TD>4.28ml</TD></TR> </TABLE></TD></TR> </TABLE> <LI>check the pH: ~0.5µl on pH-paper;</LI> <LI>check the quality and concentration (it is useful to take final dilution 1:5000 (~20µM). In this case the optical density will be in the region of A<SUB>m</SUB>~0.3 - the most accurate range for spectrophotometer).<br> Concentration is c[mM]=k<SUB>1:5000</SUB>xA<SUB>m</SUB>.</LI> </OL> <P>Quality:</P> <TABLE width="408" border="0" cellspacing="0" cellpadding="0" align="center"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="ffffff"><TD>dATP<br>pH 7.0</TD><TD>A<SUB>250</SUB>/A<SUB>260</SUB>=0.80<U>+</U>0.03<br> A<SUB>280</SUB>/A<SUB>260</SUB>=0.12<U>+</U>0.02</TD><TD>dCTP<br> pH 2.0(!)</TD><TD>A<SUB>250</SUB>/A<SUB>260</SUB>=0.45<U>+</U>0.03<br> A<SUB>280</SUB>/A<SUB>260</SUB>=2.10<U>+</U>0.15<br> A<SUB>290</SUB>/A<SUB>260</SUB>=1.60<U>+</U>0.10</TD></TR> <TR bgcolor="ffffff"><TD>dGTP<br>pH 7.5</TD><TD>A<SUB>250</SUB>/A<SUB>260</SUB>=1.18<U>+</U>0.04<br> A<SUB>280</SUB>/A<SUB>260</SUB>=0.67<U>+</U>0.03<br> A<SUB>290</SUB>/A<SUB>260</SUB>=0.28<U>+</U>0.03</TD><TD>dTTP<br> pH 7.5</TD><TD>A<SUB>250</SUB>/A<SUB>260</SUB>=0.65<U>+</U>0.03<br> A<SUB>280</SUB>/A<SUB>260</SUB>=0.73<U>+</U>0.03</TD></TR> </TABLE></TD></TR> </TABLE> <P>Concentration:</P> <TABLE width="408" border="0" cellspacing="0" cellpadding="0" align="center"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td> </td> <TD>Mw</TD><TD>A<SUB>m</SUB></TD><TD>pH</TD><TD>E</TD><TD>k (for 1:5000)</TD></TR> <TR bgcolor="ffffff"><TD>dATP[Na<SUB>2</SUB>]</TD><TD>589.2</TD><TD>259</TD><TD>7.0</TD><TD>15.2x10<SUP>3</SUP></TD><TD>328.9</TD></TR> <TR bgcolor="ffffff"><TD>dGTP[Na<SUB>3</SUB>]</TD><TD>609.2</TD><TD>253</TD><TD>7.5</TD><TD>13.7x10<SUP>3</SUP></TD><TD>365.0</TD></TR> <TR bgcolor="ffffff"><TD>dCTP[Na<SUB>3</SUB>]</TD><TD>569.1</TD><TD>280</TD><TD>2.0 (!!!)</TD><TD>13.0x10<SUP>3</SUP></TD><TD>384.6</TD></TR> <TR bgcolor="ffffff"><TD>dTTP[Na<SUB>3</SUB>]</TD><TD>584.1</TD><TD>267</TD><TD>7.5</TD><TD>9.6x10<SUP>3</SUP></TD><TD>520.8</TD></TR> <TR bgcolor="eeeeee"><td> </td> <TD> </TD><TD> </TD><TD> </TD><TD> </TD><TD> </TD></TR> <TR bgcolor="ffffff"><TD>ATP [Na<SUB>4</SUB>]</TD><TD>595.1</TD><TD>259</TD><TD> </TD><TD>15.4x10<SUP>3</SUP></TD><TD> </TD></TR> <TR bgcolor="ffffff"><TD>CTP [Na<SUB>4</SUB>]</TD><TD>571.1</TD><TD>280</TD><TD> </TD><TD>13.0x10<SUP>3</SUP></TD><TD> </TD></TR> <TR bgcolor="ffffff"><TD>GTP [Na<SUB>4</SUB>]</TD><TD>611.1</TD><TD>252</TD><TD> </TD><TD>13.7x10<SUP>3</SUP></TD><TD> </TD></TR> <TR bgcolor="ffffff"><TD>UTP [Na<SUB>4</SUB>]</TD><TD>572.1</TD><TD>262</TD><TD> </TD><TD>10.2x10<SUP>3</SUP></TD><TD> </TD></TR> </TABLE></TD></TR> </TABLE> <P>Concentration: c[M]=A<SUB>max</SUB>/E; A<SUB>max</SUB> = maximum of absorption.</P> <form><A name="a8"></A><p class=h3>EDTA</P> <P>C<SUB>10</SUB>H<SUB>14</SUB>O<SUB>8</SUB>N<SUB>2</SUB>Na<SUB>2</SUB>x2H<SUB>2</SUB>O; Mw=372.3g/M; pH = 8.0 (store at 4<SUP>o</SUP>C).</P> <TABLE width="565" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>%(w/w)</TD><TD>50 ml</TD><TD>100 ml</TD><TD>150 ml</TD><TD>250 ml</TD><TD><input type="text" size=7 onChange='clc(this,0)'></TD></TR> <TR bgcolor="ffffff"><TD>EDTA</TD><TD>0.5 M</TD><TD>372.3g/M</TD><TD>16.98</TD><TD>9.31g</TD><TD>18.62g</TD><TD>27.92g</TD><TD>46.55g</TD><TD><input type="text" size=7 onChange='clc(this,1)'></TD></TR> <TR bgcolor="ffffff"><TD>NaOH</TD><TD>~0.5 M</TD><TD>40g/M</TD><TD>1.82</TD><TD>1.014g</TD><TD>2.028g</TD><TD>3.042g</TD><TD>5.07g</TD><TD><input type="text" size=7 onChange='clc(this,2)'></TD></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD>81.19</TD><TD>44.48ml</TD><TD>88.95ml</TD><TD>133.4ml</TD><TD>222.4ml</TD><TD><input type="text" size=7 onChange='clc(this,3)'></TD></TR> </TABLE></TD></TR> </TABLE><input type="hidden" name="prm" value="100,2,ml,18.615,2,g,2.028,3,g,88.952,2,ml"></form> <form><TABLE width="565" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>10 ml</TD><TD>50 ml</TD><TD>100 ml</TD><TD>150 ml</TD><TD>250 ml</TD><TD><input type="text" size=7 onChange='clc(this,0)'></TD></TR> <TR bgcolor="ffffff"><TD>EDTA</TD><TD>0.5 M</TD><TD>372.3g/M</TD><TD>1.86g</TD><TD>9.31g</TD><TD>18.62g</TD><TD>27.92g</TD><TD>46.55g</TD><TD><input type="text" size=7 onChange='clc(this,1)'></TD></TR> <TR bgcolor="ffffff"><TD>NaOH</TD><TD>~0.5 M</TD><TD>10M</TD><TD> <u>507µl</u><br>0.674g</TD><TD><u>2.535ml</u><br>3.37g</TD><TD><u>5.07ml</u><br>6.74g</TD><TD><u>7.61ml</u><br>10.11g</TD><TD><u>12.68ml</u><br>16.85g</TD><TD> <input type="text" size=7 onChange='clc(this,2)'><br><input type="text" size=7 onChange='clc(this,3)'></TD></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD>8.42ml</TD><TD>42.12ml</TD><TD>84.24ml</TD><TD>126.4ml</TD><TD>210.6ml</TD><TD><input type="text" size=7 onChange='clc(this,4)'></TD></TR> </TABLE></TD></TR> </TABLE> <P>p~1.096g/ml</P> <li>EDTA is not soluble at acidic pH; it is necessary to add alkali gradually and to control pH;</li> <li>do not treat by DEPC.</li> <input type="hidden" name="prm" value="100,2,ml,18.615,2,g,5.07,2,ml,6.74,2,g,84.24,2,ml"></form> <form><A name="a9"></A><p class=h3>EtBr</P> <p>Ethidium bromide, C<SUB>21</SUB>H<SUB>20</SUB>N<SUB>3</SUB>Br; Mw=394.3g/M; (store at NT in the dark);</p> <TABLE width="278" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>5ml</TD><TD>10ml</TD><TD>50ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>EtBr</TD><TD>10mg/ml</TD><TD>50mg</TD><TD>100mg</TD><TD>500mg</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> </TABLE></TD></TR> </TABLE> <li>soluble in H<SUB>2</SUB>O, EtOH, chloroform;</li> <li>concerning carcinogenic properties of EtBr. The only data that we found in the literature is that in mutagenic test (on bacteria) 90µg of EtBr gave the same results as the smoke concentrate from one cigarette.</li> <input type="hidden" name="prm" value="1,2,ml,0.01,3,g"></form> <form><A name="a10"></A><p class=h3>Gelatin</P> <P>(store at 4<SUP>o</SUP>C).</P> <TABLE width="292" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>10ml</TD><TD>50ml</TD><TD>100ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>Gelatin</TD><TD>2%</TD><TD>solid</TD><TD>0.2g</TD><TD>1.0g</TD><TD>2.0g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD> </TD><TD> </TD><TD> </TD><td> </td></TR> </TABLE></TD></TR> </TABLE> <li>autoclave sterilization.</li><input type="hidden" name="prm" value="100,2,ml,2,2,g"></form> <form><A name="a11"></A><p class=h3>Glucose</P> C<sub>6</sub>H<sub>12</sub>O<sub>6</sub>xH<sub>2</sub>O, Mw=198.17g/M (store at 4<SUP>o</SUP>C)<br> <TABLE width="550" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR align="center" bgcolor="eeeeee"><td><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>%(w/w)</TD><TD>50ml</TD><TD>100ml</TD><TD>250ml</TD><TD align="center"><input type="text" size=7 onChange='clc(this,0)'></TD></TR> <TR align=center bgcolor="ffffff"><TD>Glucose</TD><TD>2M</TD><TD>198.17g/M</TD><TD>34.972</TD><TD>19.82g</TD><TD>39.63g</TD><TD>99.09g</TD><TD align="center"><input type="text" size=7 onChange='clc(this,1)'></TD></TR> <TR align=center bgcolor="ffffff"><TD>H<sub>2</sub>O</TD><TD> </TD><TD>mQ</TD><TD>65.028</TD><TD>36.85ml</TD><TD>73.70ml</TD><TD>184.24ml</TD><TD align="center"><input type="text" size=7 onChange='clc(this,2)'></TD></TR> </TABLE></TD></TR> </TABLE><input type="hidden" name="prm" value="100,2,ml,39.634,2,g,73.696,2,ml"></form> <form><TABLE width="350" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD colspan="3" align="center">1000ml/<input type="text" size=7 onChange='clc(this,0)'></TD></TR> <TR align=center bgcolor="eeeeee"><TD>198.17g/M</TD><TD>1M</TD><TD>1.5M</TD><TD>2M</TD></TR> <TR align=center bgcolor="ffffff"><TD>Glucose [g]</TD><TD>198.17<br><input type="text" size=6 onChange='clc(this,1)'></TD><TD>297.255<br><input type="text" size=6 onChange='clc(this,2)'></TD><TD>396.34<br><input type="text" size=6 onChange='clc(this,3)'></TD></TR> <TR align=center bgcolor="ffffff"><TD>H<sub>2</sub>O [ml]</TD><TD>868.23<br><input type="text" size=6 onChange='clc(this,4)'></TD><TD>802.745<br><input type="text" size=6 onChange='clc(this,5)'></TD><TD>736.96<br><input type="text" size=6 onChange='clc(this,6)'></TD></TR> <TR align=center bgcolor="ffffff"><TD>%(w/w)</TD><TD>18.583</TD><TD>27.023</TD><TD>34.972</TD></TR> <TR align=center bgcolor="ffffff"><TD>H<sub>2</sub>O %(w/w)</TD><TD>81.417</TD><TD>72.977</TD><TD>65.028</TD></TR> <TR align=center bgcolor="ffffff"><TD>p (g/ml) 20<SUP>o</SUP>C</TD><TD>1.0664</TD><TD>1.1</TD><TD>1.1333</TD></TR> </TABLE></TD></TR> </TABLE><input type="hidden" name="prm" value="100,2,ml,19.817,2,,29.7255,2,,39.634,2,,86.823,2,,80.2745,2,,73.696,2,"></form> </ul> </td></tr></table> <table width="610" border="0" cellspacing="0" cellpadding="0"><tr><td> <ul> <form><p class=h3><A name="a12"></A>Guanidine HCl (GuHCl)</P> <P>CH<SUB>5</SUB>N<SUB>3</SUB>xHCl, Mw=95.53g/M</P> <TABLE width="580" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td rowspan="2" align="center"><input type="Reset" value='Clear'></td><TD colspan="8" align="center">Molarity; 1000ml / <input type="text" size=7 onChange='clc(this,0)'></TD></tr> <TR bgcolor="eeeeee"><TD>1</TD><TD>2</TD><TD>3</TD><TD>4</TD><TD>5</TD><TD>6</TD><td>7</td><td>8</td></TR> <TR bgcolor="ffffff"><TD>GuHCl</TD><TD>95.53g<br><input type="text" size=6 onChange='clc(this,1)'></TD><TD>191.06g<br><input type="text" size=6 onChange='clc(this,2)'></TD><TD>286.59g<br><input type="text" size=6 onChange='clc(this,3)'></TD><TD>382.12g<br><input type="text" size=6 onChange='clc(this,4)'></TD><TD>477.65g<br><input type="text" size=6 onChange='clc(this,5)'></TD><TD>573.18g<br><input type="text" size=6 onChange='clc(this,6)'></TD><td>668.71g<br><input type="text" size=6 onChange='clc(this,7)'></td><td>764.24g<br><input type="text" size=6 onChange='clc(this,8)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O (mQ)</TD><TD>924.2ml<br><input type="text" size=6 onChange='clc(this,9)'></TD><TD>854ml<br><input type="text" size=6 onChange='clc(this,10)'></TD><TD>783.7ml<br><input type="text" size=6 onChange='clc(this,11)'></TD><TD>711.7ml<br><input type="text" size=6 onChange='clc(this,12)'></TD><TD>639.8ml<br><input type="text" size=6 onChange='clc(this,13)'></TD><TD>567.2ml<br><input type="text" size=6 onChange='clc(this,14)'></TD><td>494.3ml<br><input type="text" size=6 onChange='clc(this,15)'></td><td>420.7ml<br><input type="text" size=6 onChange='clc(this,16)'></td></TR> <TR bgcolor="ffffff"><TD>GuHCl %(w/w)</TD><TD>9.37</TD><TD>18.28</TD><TD>26.78</TD><TD>34.93</TD><TD>42.75</TD><TD>50.26</TD><td>57.50</td><td>64.50</td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O %(w/w)</TD><TD>90.63</TD><TD>81.72</TD><TD>73.22</TD><TD>65.07</TD><TD>57.25</TD><TD>49.74</TD><td>42.50</td><td>35.50</td></TR> <TR bgcolor="ffffff"><TD>p (g/ml)</TD><TD>1.020</TD><TD>1.045</TD><TD>1.070</TD><TD>1.094</TD><TD>1.117</TD><TD>1.140</TD><td>1.163</td><td>1.185</td></TR> </TABLE></TD></TR> </TABLE> <li>solubility: at 25<SUP>o</SUP>C - 8.54M, 5<SUP>o</SUP>C - <U>></U>8M;</li> <li>A<SUB>260</SUB>(6M in H<SUB>2</SUB>O)<0.03;</li> <li>it is possible to take the "partial density of GuHCl" as 0.763 in calculations of solutions.</li> <input type="hidden" name="prm" value="100,2,ml,9.553,2,g,19.106,2,g,28.659,2,g,38.212,2,g,47.765,2,g,57.318,2,g,66.871,2,g,76.424,2,g,92.42,2,ml,85.4,2,ml,78.37,2,ml,71.17,2,ml,63.98,2,ml,56.72,2,ml,49.43,2,ml,42.07,2,ml"></form> </ul> </td></tr></table> <table width="610" border="0" cellspacing="0" cellpadding="0"><tr><td> <ul> <form><A name="a13"></A><p class=h3>HEPES</P> <TABLE width="400" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"><TD> <TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>%(w/w)</TD><TD>1L</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD><P>C<SUB>8</SUB>H<SUB>18</SUB>N<SUB>2</SUB>O<SUB>4</SUB>S</TD><TD>1M</TD><TD>238.3g/M</TD><td>22.40</td><TD>238.3g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><td>77.60</td><TD>825.7ml</TD><td><input type="text" size=7 onChange='clc(this,2)'></td></TR> </TABLE></TD></TR></TABLE> <P>p=1.064</P><input type="hidden" name="prm" value="100,2,ml,23.83,2,g,82.57,2,ml"></form> <form><P>HEPES, 1M, 1L</P> <TABLE width="300" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><TD rowspan="2">Desired pH</TD><TD rowspan="2">KOH, 5M<br>[1000ml]</TD><td><input type="Reset" value='Clear'></td></TR> <TR bgcolor="eeeeee"><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>5.25</TD><TD>0ml</TD><td>0ml</td></TR> <TR bgcolor="ffffff"><TD>5.35</TD><TD>0.5ml</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>5.75</TD><TD>3.5ml</TD><td><input type="text" size=7 onChange='clc(this,2)'></td></TR> <TR bgcolor="ffffff"><TD>6.03</TD><TD>7ml</TD><td><input type="text" size=7 onChange='clc(this,3)'></td></TR> <TR bgcolor="ffffff"><TD>6.24</TD><TD>12ml</TD><td><input type="text" size=7 onChange='clc(this,4)'></td></TR> <TR bgcolor="ffffff"><TD>6.59</TD><TD>22ml</TD><td><input type="text" size=7 onChange='clc(this,5)'></td></TR> <TR bgcolor="ffffff"><TD>6.71</TD><TD>32ml</TD><td><input type="text" size=7 onChange='clc(this,6)'></td></TR> <TR bgcolor="ffffff"><TD>6.88</TD><TD>45ml</TD><td><input type="text" size=7 onChange='clc(this,7)'></td></TR> <TR bgcolor="ffffff"><TD>7.00</TD><TD>50ml</TD><td><input type="text" size=7 onChange='clc(this,8)'></td></TR> <TR bgcolor="ffffff"><TD>7.10</TD><TD>60ml</TD><td><input type="text" size=7 onChange='clc(this,9)'></td></TR> <TR bgcolor="ffffff"><TD>7.25</TD><TD>80ml</TD><td><input type="text" size=7 onChange='clc(this,10)'></td></TR> <TR bgcolor="ffffff"><TD>7.37</TD><TD>92.5ml</TD><td><input type="text" size=7 onChange='clc(this,11)'></td></TR> </TABLE></TD></TR> </TABLE><input type="hidden" name="prm" value="100,2,ml,0.05,3,ml,0.35,3,ml,0.7,3,ml,1.2,2,ml,2.2,2,ml,3.2,2,ml,4.5,2,ml,5,2,ml,6,2,ml,8,2,ml,9.25,2,ml"></form> <form><A name="a14"></A><p class=h3>Imidazol</P> <P>C<SUB>3</SUB>H<SUB>4</SUB>N<SUB>2</SUB>, (store at 4<SUP>o</SUP>C):</P> <TABLE width="400" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>Stock</TD><TD>50ml</TD><TD>100ml</TD><td><input type="text" size=7 onChange='clc(this,0)'></td></TR> <TR bgcolor="ffffff"><TD>Imidazole</TD><TD>2M</TD><TD>68.1g/M</TD><TD>6.81g</TD><TD>13.62g</TD><td><input type="text" size=7 onChange='clc(this,1)'></td></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD> </TD><TD>mQ</TD><TD> </TD><TD> </TD><td> </td></TR> </TABLE></TD></TR> </TABLE><input type="hidden" name="prm" value="100,2,ml,13.62,2,g"></form> <A name="a15"></A><p class=h3>Paraformaldehyde</P> <P>PFA (paraformaldehyde) 37% (for histochemistry it should be freshly prepared).</P> <P>1. mix in the screw-cap tube:</P> <P class=ls>PFA (solid) = 0.37g,<br> H<SUB>2</SUB>O = 1.0ml<br> NaOH (1N) = 14µl;</P> <P>2. solubilize in the boiling water bath (to heat ~1-3'; until pH will drop to ~7.0).</P> <form><A name="a16"></A><p class=h3>PEG</P> <P>H(OCH<SUB>2</SUB>CH<SUB>2</SUB>)<SUB>n</SUB>OH; (store at 4<SUP>o</SUP>C).</P> <TABLE width="507" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"><input type="Reset" value='Clear'></td><TD>Conc.</TD><TD>%(w/w)</TD><TD>10ml</TD><TD>50ml</TD><TD>100ml</TD><TD>150ml</TD><TD>200ml</TD><TD><input type="text" size=7 onChange='clc(this,0)'></TD></TR> <TR bgcolor="ffffff"><TD>PEG6000</TD><TD>40%</TD><TD>37.21</TD><TD>4.0g</TD><TD>20g</TD><TD>40g</TD><TD>60g</TD><TD>80g</TD><TD><input type="text" size=7 onChange='clc(this,1)'></TD></TR> <TR bgcolor="ffffff"><TD>H<SUB>2</SUB>O</TD><TD>mQ</TD><TD>62.79</TD><TD>6.75g</TD><TD>33.75g</TD><TD>67.5g</TD><TD>101.25g</TD><TD>135.0g</TD><TD><input type="text" size=7 onChange='clc(this,2)'></TD></TR> </TABLE></TD></TR> </TABLE> <P>p=1.075.</P><input type="hidden" name="prm" value="100,2,ml,40,2,g,67.5,2,g"></form> <form><A name="a17"></A><p class=h3>PMSF</P> <P>(store at -20<SUP>o</SUP>C)</P> <TABLE width="450" border="0" cellspacing="0" cellpadding="0"> <TR bgcolor="000000"> <TD><TABLE width="100%" border="0" cellspacing="1" cellpadding="2px"> <TR bgcolor="eeeeee"><td align="center"