https://eln.excellgen.com/index.php?action=history&feed=atom&title=PCR_from_Plasmid_and_Genomic_DNAPCR from Plasmid and Genomic DNA - Revision history2026-10-18T07:50:38ZRevision history for this page on the wikiMediaWiki 1.44.0https://eln.excellgen.com/index.php?title=PCR_from_Plasmid_and_Genomic_DNA&diff=16141&oldid=prevPaul.xu at 15:54, 23 June 20172017-06-23T15:54:49Z<p></p>
<p><b>New page</b></p><div>Must use high fiderity polymerase such as Phusion DNA Polymerase<br />
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== Genomic DNA ==<br />
<br />
25 ~35 cycles<br />
<br />
25 or 50 ul reaction<br />
<br />
5 ~10 ul genomic DNA, at least 100 copies&nbsp;<br />
<br />
0.5~1 ul F 10 uM primer<br />
<br />
0.5~1 ul R 10 uM primer<br />
<br />
0.5~1 ul 10 mM dNTP<br />
<br />
5 or 10 ul &nbsp;5X Phusion or Q5 Buffer<br />
<br />
0.5 or 1 ul Phusion<br />
<br />
<br />
<br />
Run 15 ul + 5 ul 6X GRGreen Loading Buffer on gel.<br />
<br />
Clone PCR product using [[Blunt-end_PCR_Cloning|blunt-end cloning protocol]]<br />
<br />
== Plasmid DNA ==<br />
<br />
25 &nbsp; cycles<br />
<br />
25 or 50 ul reaction<br />
<br />
0.5 ul plasmid &nbsp;DNA, at least 100 copies&nbsp;<br />
<br />
0.5~1 ul F 10 uM primer<br />
<br />
0.5~1 ul R 10 uM&nbsp;primer<br />
<br />
0.5~1 ul 10 mM dNTP<br />
<br />
5 or 10 ul &nbsp;5X Phusion or Q5 Buffer<br />
<br />
0.5 or 1 ul Phusion<br />
<br />
<br />
<br />
Run 15 ul + 5 ul 5X GRGreen Loading Buffer on gel.<br />
<br />
Clone PCR product using [[Blunt-end_PCR_Cloning|blunt-end cloning protocol]]<br />
<br />
'''PCR Positive Control'''<br />
<br />
Usually you need to run at least one positive control --> your PCR is working<br />
<br />
and one negative control.-->identify false positive<br />
[[Category:Protocols]]</div>Paul.xu