Team:Warsaw/Calendar-Main/7 October 2008

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<h3>Preparation of <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + _alpha</h3>
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<h4>Michał K.</h4>
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<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmid from culture inoculated on previous day (<a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + _alpha).</li>
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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 plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - no proper clones found.</li></ol>
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<h3>Preparation of BioBricks</h3>
<h3>Preparation of BioBricks</h3>
<h4>Michał K.</h4>
<h4>Michał K.</h4>

Revision as of 21:46, 26 October 2008

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Preparation of pSB2K3 + _alpha

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day (pSB2K3 + _alpha).
  2. Control digest of isolated plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - no proper clones found.

Preparation of BioBricks

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day (pSB2K3 + _alpha, pSB2K3 + _omega and pACYC177 + OmpA_omega_).
  2. Control digest of isolated plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - no proper clones found.
  3. Overnight ligation of DNA fragments: pSB1A3+ AID.

Piotr

  1. Transformation of TOP10 with ligations pSB2K3 + pLac_OmpA_omega (without EcoRI site) and pET15b+OmpA_omega without XbaI.
  2. Plating: pSB2K3 + pLac_OmpA_omega (without EcoRI site) on LB with kanamycin and pET15b+OmpA_omega without XbaI on LB with ampicillin.
  3. Inoculation of few more colonies which grown on plates with three separate transformations: pSB2K3 + _alpha, pSB2K3 + _omega and pACYC177 + OmpA_omega_ to liquid LB + kanamycin.
  4. Digest of pMPMT5+AID with EcoRI (EcoRI buffer). DNA ends blunting with Klenow fragment.
  5. Overnight ligation of pMPMT5+AID without EcoRI site.

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