https://eln.excellgen.com/index.php?action=history&feed=atom&title=RT-PCR 2026-10-18T04:56:46Z Revision history for this page on the wiki MediaWiki 1.44.0 https://eln.excellgen.com/index.php?title=RT-PCR&diff=956&oldid=prev 2011-06-05T02:12:12Z

<p></p> <p><b>New page</b></p><div>&lt;u&gt;&#039;&#039;&#039;RT-PCR for lentivirus tittering&#039;&#039;&#039;&lt;/u&gt;&lt;br&gt;&#039;&#039;&#039;Virus treatment:&#039;&#039;&#039;&lt;br&gt;1. Add 12 ul virus containing medium to 108 ul lysis buffer (10 mM glycine, 0.5% Triton X-100, pH 2.5);<br /> <br /> &lt;br&gt;2. incubate at RT for 10 min;&lt;br&gt;3. Add 2 ul 1M Tris pH 8.0 to adjust pH to about 7.0;&lt;br&gt;4. Add 1 ul 10mg/ml DNase;&lt;br&gt;5. Incubate at RT for 30 min;&lt;br&gt;6. Incubate at 75 oC for 10min to denature DNase;&lt;br&gt;7. Use 2 ul to do RT-PCR.&lt;br&gt;<br /> <br /> &#039;&#039;&#039;RT-PCR:&#039;&#039;&#039;&lt;br&gt;2 ul virus sample treated above;&lt;br&gt;1 ul 10 uM WPRE-F&lt;br&gt;1 ul 10 uM WPRE-R&lt;br&gt;12.5 ul 2 X Mastermix&lt;br&gt;8.5 ul ddH2O&lt;br&gt;Total 25 ul.<br /> <br /> &lt;br&gt;Use ABI7000 to do the RT-PCR.&lt;br&gt;1. 50°C 2 min;&lt;br&gt;2. 95°C 10 min;&lt;br&gt;3. 95°C 15 S;&lt;br&gt;4. 60°C 1 min;&lt;br&gt;5. Repeat step 3 and 4 for another 39 cycles.&lt;br&gt;&lt;br&gt; --[[User:Ztao|Ztao]] 09:53, 12 April 2011 (EST) <br /> <br /> [[Category:Protocols]]</div>

Paul.xu