Team:Warsaw/Calendar-Main/7 October 2008

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<li> Planting: (3kb??????) + pLac_OmpA_omega (without EcoRI site) on LB with kanamycin and pET15b+OmpA_omega without XbaI on LB with ampicillin.</li>
<li> Planting: (3kb??????) + pLac_OmpA_omega (without EcoRI site) on LB with kanamycin and pET15b+OmpA_omega without XbaI on LB with ampicillin.</li>
<li>Inoculation of few more colonies which grown on plates with three separate transformations: (3kb??????) + _alpha, (3kb????) + _omega and pACYC177 + OmpA_omega_ to liquid LB + kanamycin. </li>
<li>Inoculation of few more colonies which grown on plates with three separate transformations: (3kb??????) + _alpha, (3kb????) + _omega and pACYC177 + OmpA_omega_ to liquid LB + kanamycin. </li>
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<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#digest">Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pMPM-T5-AID>pMPMT5+AID</a> with EcoRI (EcoRI buffer). DNA ends <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#blunting">blunting</a> with Klenow fragment. </li>
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<li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pMPM-T5-AID>pMPMT5+AID</a> without EcoRI site.</li>
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</ol>

Revision as of 11:33, 25 October 2008

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Preparation of BioBricks

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day ((3kb??????) + _alpha, (3kb????) + _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: RFP(????)+ AID.

Piotr

  1. Transformation of TOP10 with ligations (3kb??????) + pLac_OmpA_omega (without EcoRI site) and pET15b+OmpA_omega without XbaI.
  2. Planting: (3kb??????) + 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: (3kb??????) + _alpha, (3kb????) + _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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