Team:Warsaw/Calendar-Main/25 June 2008

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<h4>Piotr, Weronika</h4>
<h4>Piotr, Weronika</h4>
<ol>
<ol>
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<li>Inoculation of 10ml LB with E.coli TOP10 carrying pZC</li>
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<li>Inoculation of 10ml LB with E.coli TOP10 carrying pZC.</li>
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<li>Isolation and chemotransformation with <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>Isolation and chemotransformation with <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>Plating of chemotransformants on LB+ampicillin
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<li>Plating of chemotransformants on LB+ampicillin.
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</li></ol>
</li></ol>
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<h3>Preparation of constructs with OmpA protein fusions</h3>
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<h3>Preparation of constructs: OmpA_alpha and OmpA_omega</h3>
<h4>Michał K.</h4>
<h4>Michał K.</h4>
<p>Repetition of PCRs and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out purifications</a> from <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/16_June_2008">16 June.</a>
<p>Repetition of PCRs and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out purifications</a> from <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/16_June_2008">16 June.</a>

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

Piotr, Weronika

  1. Inoculation of 10ml LB with E.coli TOP10 carrying pZC.
  2. Isolation and chemotransformation with pSB1A2 standard plasmids carrying parts: BBa_E0840(GFP genetrator) and BBa_J04450 (RFP generator).
  3. Plating of chemotransformants on LB+ampicillin.

Preparation of constructs: OmpA_alpha and OmpA_omega

Michał K.

Repetition of PCRs and gel-out purifications from 16 June.

Preparation of alpha-A and omega-A fusions

Michał L. Marcin and Ewa

  1. We've run gradient PCR to obtain Alpha-link and link-A.
    • DNA template: pDRIVE-TAPTAG
    • Taq polymerase
    • buffer suplemented with (NH4)2SO4
    • MgCl2 concentration gradient (from 1 to 4 mM)
    • annealing temp 55°C
    • elongation time 1 - 2 minutes
  2. We have successfully amplified Alpha-link and link-A (Fig. 1). Now it's time for PCL (Polymerase Chain Ligation) to create fusions Alpha-A and Omega-A.
Fig. 1. PCR product - link-A.

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