G.tigrina Hox gene DthoxC insertion into prokaryote E.coli / (by UNIamCloning)/2011/10/17: Difference between revisions

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==PCR Check of Sample Gt.5-2, Ambiguous COI Primers (New Positive Control) and Newly Diluted DthoxC primers==
==PCR Check of Sample Gt.5-2, Ambiguous COI Primers (New Positive Control) and Newly Diluted DthoxC primers==
* Today we ran an electrophoresis gel on last week's final PCR amplification for the planarian gene DthoxC.  (It was decided that if this PCR failed, the project would be abandoned in favor of something more likely to succeed.)  We decided to make two sets of PCR tubes - some using the primers we designed for amplifying DthoxC, only with a slightly stronger concentration after dilution - as well as a set of primers designed to amplify the mitochondrial COI gene, just so that we could see if the problem was in our DNA extract or with our primer design.
* Today we ran an electrophoresis gel on last week's final PCR amplification for the planarian gene DthoxC.  (It was decided that if this PCR failed, the project would be abandoned in favor of something more likely to succeed.)  We decided to make two sets of PCR tubes - some using the primers we designed for amplifying DthoxC, only with a slightly stronger concentration after dilution - as well as a set of primers designed to amplify the mitochondrial COI gene, just so that we could see if the problem was in our DNA extract or with our primer design.
*Result:  The gel revealed that the COI primers worked marvelously in amplifying the COI gene, meaning that there was not fault in our DNA extractions.  The DthoxC primer samples showed multiple, faint bands on the gel... probably primer dimer effects/hairpins/etc.  So, it was simply a primer design that did not work the way we wanted it to.




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PCR Check of Sample Gt.5-2, Ambiguous COI Primers (New Positive Control) and Newly Diluted DthoxC primers

  • Today we ran an electrophoresis gel on last week's final PCR amplification for the planarian gene DthoxC. (It was decided that if this PCR failed, the project would be abandoned in favor of something more likely to succeed.) We decided to make two sets of PCR tubes - some using the primers we designed for amplifying DthoxC, only with a slightly stronger concentration after dilution - as well as a set of primers designed to amplify the mitochondrial COI gene, just so that we could see if the problem was in our DNA extract or with our primer design.
  • Result: The gel revealed that the COI primers worked marvelously in amplifying the COI gene, meaning that there was not fault in our DNA extractions. The DthoxC primer samples showed multiple, faint bands on the gel... probably primer dimer effects/hairpins/etc. So, it was simply a primer design that did not work the way we wanted it to.


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