Team:Warsaw/Calendar-Main/27 August 2008

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<h3>Cloning of protein A DNA to pET15b+Omp-alfa plasmid</h3><h4>Michał K.</h4>
<h3>Cloning of protein A DNA to pET15b+Omp-alfa plasmid</h3><h4>Michał K.</h4>
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<ol><li> Digestion of pGeneart+A and pET15b+Omp-alfa with NdeI and NotI (Orange buffer), pET15b was also dephosphorylated.</li>
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<ol><li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of pGeneart+A and pET15b+Omp-alfa with NdeI and NotI (Orange buffer), pET15b was also <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing"> dephosphorylated</a>. </li> .</li>
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<li> Gel electrophoresis of digested plasmids and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (400 bp for A lane and 6200 bp for pET15b lane).  </li>
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<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">Ligation</a> of pET15b+alfa and A. </li>
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<li> Transformation of <i>E. coli</i> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a>. </li>
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<li> Transformants plating on LB + ampicillin.</li>
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Cloning of protein Z DNA to pET15b+Omp-alfa plasmid

Antoni

Transformant colonies inoculated to liquid LB with ampicillin.

Cloning of protein A DNA to pET15b+Omp-alfa plasmid

Michał K.

  1. Digest of pGeneart+A and pET15b+Omp-alfa with NdeI and NotI (Orange buffer), pET15b was also dephosphorylated.
  2. .
  3. Gel electrophoresis of digested plasmids and gel-out of proper bands (400 bp for A lane and 6200 bp for pET15b lane).
  4. Ligation of pET15b+alfa and A.
  5. Transformation of E. coli TOP10.
  6. Transformants plating on LB + ampicillin.