<div style="margin: 12px 0px; font-family: arial; color: #333333; background: #ffffff; border: solid 4px #e5e5e5; width: 100%; clear: left;"><div class="CM_CTB_Content_Wrap" style="margin: 0px; padding: 0px;background-color: #ffffff;"><div style="border-bottom: solid 1px #dcdcdc; white-space: nowrap; margin-bottom: 8px; background-color: #eeeeee ;background-image: url(http://clipmarks.com/images/source-bg.gif); background-repeat: repeat-x; height: 24px; line-height: 24px; vertical-align: middle; padding-bottom: 4px; color: #666666; font-size: 10px;" ><a href="http://clipmarks.com/clip-to-blog/" title="see clips that are hot right now"><img src="http://content.clipmarks.com/blog_embed/8b61672d-a0d1-40af-a2c7-a2ca842ce441/DECFC5FC-095A-4F25-BC0C-920885BD1CD6/" alt="" width="19" height="19" border="0" style="vertical-align: middle; margin: 0px 4px; display: inline; border: none; float:none;" /></a>clipped from <a title="http://www.fermentas.com/techinfo/electrophoresis/rnaloading.htm" href="http://www.fermentas.com/techinfo/electrophoresis/rnaloading.htm" style="font-size: 11px;">www.fermentas.com</a></div><blockquote style="text-align: left; padding: 0px 8px; margin: 4px 0px 8px 0px; background: transparent; border: none;" cite="http://www.fermentas.com/techinfo/electrophoresis/rnaloading.htm"><DIV class="bsup"><B>Denaturing <A name="polyacrylamide">polyacrylamide</A>/urea gels in TBE buffer</B></DIV> <OL> <li style="margin-left:16px;padding-left: 0px;">Prepare 20 ml of a 5% polyacrylamide gel containing 7 M urea by adding:<TABLE cellspacing="0" cellpadding="3" bordercolor="#bdf2f2" border="1" bgcolor="#ffffff" id="table1"><TBODY><TR><TD>47.5% acrylamide: 2.5% bis-acrylamide solution</TD><TD>2 ml </TD></TR><TR><TD>10 M urea</TD><TD>14 ml</TD></TR><TR><TD><A href="http://www.fermentas.com/techinfo/electrophoresis/../../catalog/electrophoresis/electrophorbuff.htm">10X TBE buffer</A></TD><TD>2 ml </TD></TR><TR><TD>10% freshly prepared ammonium persulfate</TD><TD>0.2 ml</TD></TR><TR><TD>deionized water</TD><TD>1.8 ml</TD></TR></TBODY></TABLE></LI> <li style="margin-left:16px;padding-left: 0px;">Mix and add 10 µl TEMED</LI> <li style="margin-left:16px;padding-left: 0px;">Mix again and pour the gel carefully avoiding the formation of air bubbles.</LI> <li style="margin-left:16px;padding-left: 0px;">Insert the comb into the acrylamide and allow the gel to polymerize for at least 1 hour.</LI> <li style="margin-left:16px;padding-left: 0px;">Fill the electrophoresis apparatus with 1X TBE buffer</LI> <li style="margin-left:16px;padding-left: 0px;">Heat the RNA samples (premixed with equal volume of <A href="http://www.fermentas.com/techinfo/electrophoresis/../../catalog/electrophoresis/2xloadingdye.htm">2X RNA Loading Dye</A>) and ladder at 70°C for 10 min, and chill on ice for 3 minutes. Load onto the gel.</LI> <li style="margin-left:16px;padding-left: 0px;">Run electrophoresis at 8 V/cm for about 1 hour.</LI> <li style="margin-left:16px;padding-left: 0px;">Soak the gel for about 15 minutes in 1X TBE to remove urea prior to staining.</LI> <li style="margin-left:16px;padding-left: 0px;">Stain the gel in 0.5 µg/ml ethidium bromide in 1X TBE solution for 15 min.</LI></OL></blockquote></div><div style="margin: 0px 6px 6px 4px;"><table style="font-size: 11px;border-spacing: 0px;padding: 0px;" cellpadding="0" cellspacing="0" width="100%"><tr><td style="background:transparent;border-width:0px;padding:0px;">&nbsp;</td><td align="right" style="background:transparent;border-width:0px;padding:0px;width:107px" width="107"><a href="http://clipmarks.com/share/DECFC5FC-095A-4F25-BC0C-920885BD1CD6/blog/" title="blog or email this clip"><img src="http://content6.clipmarks.com/images/c2b-foot.png" border="0" alt="blog it" width="107" height="17" style="border-width:0px;padding:0px;margin:0px;" /></a></td></tr></table></div></div>

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