Team:Warsaw/Calendar-Main/18 July 2008

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<li> Isolation of plasmids from cultures inocluated on previous day (pGeneart+A and pET15b+OmpA-alfa). </li>
<li> Isolation of plasmids from cultures inocluated on previous day (pGeneart+A and pET15b+OmpA-alfa). </li>
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<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> pET15b+OmpA_alpha and pGeneart+A with NdeI and NotI, pET15b+OmpA_alpha<a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing">dephosphorylation with CIAP</a> </li> </li>
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<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> pET15b+OmpA_alpha and pGeneart+A with NdeI and NotI, pET15b+OmpA_alpha<a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing"> dephosphorylation with CIAP</a> </li> </li>
<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  </li>
<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  </li>
<li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of pET15b+alfa and A </li>
<li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of pET15b+alfa and A </li>

Revision as of 06:48, 2 October 2008

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Cloning of protein A DNA to OmpA constructs

  1. Colony PCR on colonies from plates with transformations pACYC177+OmpA_A_omega and pACYC177+OmpA_A_alpha. Primers used: AL+SacI and AP+NotI
  2. Confirmed transformant colonies (found only on pACYC177+OmpA_A_alpha plate) inoculated to liquid LB with kanamycin.

Paweł

  1. Ligation transformed into TOP10 strain and plated on LB+kanamycin


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

  1. Isolation of plasmids from cultures inocluated on previous day (pGeneart+A and pET15b+OmpA-alfa).
  2. Digest pET15b+OmpA_alpha and pGeneart+A with NdeI and NotI, pET15b+OmpA_alpha dephosphorylation with CIAP
  3. Gel electrophoresis of digested plasmids and gel-out of proper bands
  4. Overnight ligation of pET15b+alfa and A

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