Team:Warsaw/Calendar-Main/1 July 2008

From 2008.igem.org

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<h3>Change of the reporter from pZC with B-galactosidaze to GFP or RFP</h3>
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<h3>Change of the reporter from <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pZC320>pZC320</a> with B-galactosidase to GFP or RFP</h3>
<h4>Piotr, Weronika</h4>
<h4>Piotr, Weronika</h4>
<ol>
<ol>
<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of plasmids from transformants.</li>
<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of plasmids from transformants.</li>
<li>Control <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest"> digest</a> of plasmids with NotI.</li>
<li>Control <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest"> digest</a> of plasmids with NotI.</li>
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<li>Gel electrophoresis of digested DNA - there where no proper plasmids</li>
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<li>Gel electrophoresis of digested DNA - there where no proper plasmids.</li>
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<li>Isolation of pZC</li>
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<li>Isolation of <A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pZC320>pZC320</a>.</li>
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<li>Isolation of nazwa (Gfp genetrator), nazwa (RFP generator)</li>
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<li>Isolation of <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A2>pSB1A2</a> standard plasmids carrying parts: <A href=http://partsregistry.org/wiki/index.php?title=Part:BBa_E0840>BBa_E0840</a>(GFP genetrator) and <a href=http://partsregistry.org/wiki/index.php?title=Part:BBa_J04450>BBa_J04450</a> (RFP generator).</li>
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<li>Owernight digest of nazwa (Gfp genetrator), nazwa (RFP generator) with NotI</li>
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<li>Overnight digest of <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A2>pSB1A2</a> standard plasmids carrying parts: <A href=http://partsregistry.org/wiki/index.php?title=Part:BBa_E0840>BBa_E0840</a>(GFP genetrator) and <a href=http://partsregistry.org/wiki/index.php?title=Part:BBa_J04450>BBa_J04450</a> (RFP generator) with NotI.</li>
<li>
<li>
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Owernight digest of pZC with NotI and defosforylation with CIAP
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Overnight digest of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pZC320>pZC320</a> with NotI and <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing>dephosphorylation</a> with CIAP.
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</li>
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</li></ol>
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<h3>Cloning of alpha-A and omega-A fusions on <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2B>pKS</a></h3>
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<h3>Cloning alpha-A and omega-A fusions on <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pKSII%2B>pKS</a></h3>
<h4>Michał L., Ewa, Marcin</h4>
<h4>Michał L., Ewa, Marcin</h4>
<ol>
<ol>
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<h3>Preparation of constructs with OmpA protein fusions</h3><h4>Paweł</h4>
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<h3>Preparation of constructs on pET15b: OmpA_alpha and OmpA_omega</h3><h4>Paweł</h4>
<p><ol>
<p><ol>
<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of <a href=http://www.emdbiosciences.com/docs/docs/PROT/TB045.pdf>pET15b</a> plasmid.</li>
<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of <a href=http://www.emdbiosciences.com/docs/docs/PROT/TB045.pdf>pET15b</a> plasmid.</li>

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Change of the reporter from pZC320 with B-galactosidase to GFP or RFP

Piotr, Weronika

  1. Isolation of plasmids from transformants.
  2. Control digest of plasmids with NotI.
  3. Gel electrophoresis of digested DNA - there where no proper plasmids.
  4. Isolation of pZC320.
  5. Isolation of pSB1A2 standard plasmids carrying parts: BBa_E0840(GFP genetrator) and BBa_J04450 (RFP generator).
  6. Overnight digest of pSB1A2 standard plasmids carrying parts: BBa_E0840(GFP genetrator) and BBa_J04450 (RFP generator) with NotI.
  7. Overnight digest of pZC320 with NotI and dephosphorylation with CIAP.

Cloning alpha-A and omega-A fusions on pKS

Michał L., Ewa, Marcin

  1. Isolation of plasmids from transformants.
  2. Control digest of plasmids with SacI+NotI (BamHI buffer).
  3. Gel electrophoresis of digested DNA.

We have successfully cloned A-alpha and A-omega fusions on pKS vector. The DNA will be now sequenced to ensure that our constructs are correct.

Preparation of constructs on pET15b: OmpA_alpha and OmpA_omega

Paweł

  1. Isolation of pET15b plasmid.
  2. Overnight digest of pET15b plasmid with NdeI and BamHI (Tango 2x buffer), dephosphorylation with CIAP.

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