Team:Warsaw/Calendar-Main/24 June 2008

From 2008.igem.org

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<p>Still no success. We need to run gradient PCR to find optimal reaction conditions.</p>  
<p>Still no success. We need to run gradient PCR to find optimal reaction conditions.</p>  
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<h3>Cloning of protein Z DNA to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b-OmpA-alpha</a> in place of OmpA</h3>
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<h4>Paweł</h4>
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<li>Control <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>digest</a> of isolated plasmids with NdeI and NotI. Two positives obtained (proper band: ~200 bp visible).</li>
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<li>One of positive plasmids transformed into <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> and plated on LB+amp, overnight, for further isolation of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2Bhis%2BZ%2Bomega>pET15b+Z+omega</a> vector.</li>
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<li>Gel electrophoresis (<a href="https://2008.igem.org/wiki/index.php?title=Team:Warsaw/Calendar-Main/24_June_2008#fig1">Fig. 1</a>, no proper clones found).</li>
<li>Gel electrophoresis (<a href="https://2008.igem.org/wiki/index.php?title=Team:Warsaw/Calendar-Main/24_June_2008#fig1">Fig. 1</a>, no proper clones found).</li>
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Revision as of 20:17, 19 October 2008

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Preparation of alpha-A and omega-A fusions

Michał L., Ewa, Marcin

Still no success. We need to run gradient PCR to find optimal reaction conditions.

Cloning of protein Z DNA to pET15b-OmpA-alpha in place of OmpA

Paweł

  1. Control digest of isolated plasmids with NdeI and NotI. Two positives obtained (proper band: ~200 bp visible).
  2. One of positive plasmids transformed into TOP10 and plated on LB+amp, overnight, for further isolation of pET15b+Z+omega vector.

Preparation of constructs with OmpA protein fusions

Michał K.

  1. PCR on isolated plasmids with OmpaL_N and OmpaP_link primers.
  2. Gel electrophoresis (Fig. 1, no proper clones found).

Fig. 1. PCR product - searching for fusions with OmpA protein. Lower lanes - OmpA_alpha, upper - OmpA_omega.

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