Team:Warsaw/Calendar-Main/26 June 2008

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<h3>Preparation of pMPMT5-AID+AIDT7 construct </h3>
<h3>Preparation of pMPMT5-AID+AIDT7 construct </h3>
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<ol><li>Transformation of E. coli <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a>  strain with <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> from previous day </li>
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<ol><li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform>Transformation</a> of E. coli <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a>  strain with <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> from previous day </li>
<li>Plating transformants on LB + tetracycline</li>
<li>Plating transformants on LB + tetracycline</li>

Revision as of 00:37, 5 October 2008

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Preparation of pMPMT5-AID+AIDT7 construct

  1. TOP10 strain with ligation from previous day
  2. Plating transformants on LB + tetracycline

    3. Polymerase Chain Ligation (PCL)

    ProductTemplatesPrimersProduct length
    Alpha-A fusionAlpha-linker + linker-AAlphaL+SacI + AP+NotI 1500 bp
    Omega-A fusionOmega-linker + linker-AOmegaL+SacI + AP+NotI 1440 bp

    PCL program for fusion A with alpha and omega

    TemperatureTime
    94°C4:00
    94°C0:3028 cycles
    48°C-62°C0:45
    72°C2:30
    72°C10:00
    4°Cinfinite

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