Team:Warsaw/Calendar-Main/24 July 2008

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<p>1. Separate transformant colonies (tranformation from previous day) inoculated to liquid LB with kanamycin.
 
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<p>Paweł: <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Plasmid DNA isolation</a> from overnight grown cultures. Restriction analysis (BamHI+NdeI) of 5 of them</p>
 
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<p>Results: 2 of analysed plasmids contain OmpA+omega (~1350 bp band)</p>
 
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<p>First of positive plasmids and Z (in GeneArt vector) digest with NdeI and NotI (overnight)<?p>
 
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<h3>Cloning of protein Z DNA to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-omega>pET15b-OmpA-omega</a> in place of OmpA</h3><h4>Paweł</h4>
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<ol><li>Results of ligation: 6 colonies grown.</li>
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<li>Each colony cultured overnight in LB + ampicillin.</li></ol>
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<h3>Cloning of protein A DNA to OmpA constructs</h3><h4>Michał K.</h4>
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<li>Colony PCR on colonies from plates with transformations <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-A-omega>pACYC177+OmpA_A_omega</a>. Primers used:
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<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AL+SacI">AL+SacI</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AP+NotI">AP+NotI</a>.</li><li> Gel electrophoresis.</li>
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<li>Confirmed transformant colonies inoculated to liquid LB with kanamycin. </li></ol>
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<h3>Preparation of chemocompetent bacteria</h3>
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<h4>Antoni</h4>
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<p>Preparation of <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemocompetent">chemocompetent</a> bacteria <i>E. coli</i> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htmtop10">TOP10</a>.</p>
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Latest revision as of 21:22, 29 October 2008

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Cloning of protein Z DNA to pET15b-OmpA-omega in place of OmpA

Paweł

  1. Results of ligation: 6 colonies grown.
  2. Each colony cultured overnight in LB + ampicillin.

Cloning of protein A DNA to OmpA constructs

Michał K.

  1. Colony PCR on colonies from plates with transformations pACYC177+OmpA_A_omega. Primers used: AL+SacI and AP+NotI.
  2. Gel electrophoresis.
  3. Confirmed transformant colonies inoculated to liquid LB with kanamycin.

Preparation of chemocompetent bacteria

Antoni

Preparation of chemocompetent bacteria E. coli TOP10.

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