Team:Warsaw/Calendar-Main/24 June 2008

From 2008.igem.org

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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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<h3>Preparation of constructs: OmpA_alpha and OmpA_omega</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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<h3>Preparation of constructs with OmpA protein fusions</h3>
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<h4>Michał K.</h4>
<h4>Michał K.</h4>
<ol><li> PCR on isolated plasmids with <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpaL_N">OmpaL_N</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpaP_link">OmpaP_link</a>
<ol><li> PCR on isolated plasmids with <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpaL_N">OmpaL_N</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpaP_link">OmpaP_link</a>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/7/78/Omp_colony_PCR_WAW.jpg" width=400/></a>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/7/78/Omp_colony_PCR_WAW.jpg" width=350/></a>
<var><b>Fig. 1.</b> PCR product - searching for fusions with OmpA protein. Lower lanes - OmpA_alpha, upper - OmpA_omega.</var>
<var><b>Fig. 1.</b> PCR product - searching for fusions with OmpA protein. Lower lanes - OmpA_alpha, upper - OmpA_omega.</var>

Latest revision as of 16:53, 28 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.

Preparation of constructs: OmpA_alpha and OmpA_omega

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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