Team:Warsaw/Calendar-Main/14 October 2008

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<h3>Preparation of BioBricks</h3>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103009>linker_alpha (BBa_K103009)</a></h3>
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<h4>Piotr</h4>
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<ol>
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<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> with overnight ligation - <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + <a href=http://partsregistry.org/Part:BBa_K103009>linker_alpha (BBa_K103009)</a>.</li>
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<li> Tranformants plating on LB with kanamycin. </li>
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</ol>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103016>OmpA-linker-omega-linker (BBa_K103016)</a></h3>
<h4>Michał K.</h4>
<h4>Michał K.</h4>
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<ol><li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmid from culture inoculated on previous day (5kb (??????) + pT7_omega_link, RFP(?????) + pT7_alpha and 3kb (??????) + OmpA_omega_link).</li>
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<ol><li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmid from culture inoculated on previous day - <a href=http://partsregistry.org/Part:BBa_I739204>pACYC177</a> + <a href=http://partsregistry.org/Part:BBa_K103016>OmpA-linker-omega-linker (BBa_K103016)</a>.</li>
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<li>Control <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>digest</a> of isolated plasmids (5kb (??????) + pT7_omega_link and 3kb (??????) + OmpA_omega_link) with EcoRI and PstI (Orange buffer). Gel electrophoresis - still no proper clones found for 3kb (??????) + OmpA_omega_link ligation but we found good clones for (5kb (??????) + pT7_omega_link.</li></ol>
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<li>Control <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>digest</a> of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - still no proper clones found. <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/14_October_2008#fig1">Fig. 1</a>.</li></ol>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/3/37/Traw_18_10_2008.jpg"></a>
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<var><b>Fig. 1.</b>EcoRI/PstI digests of isolated plasmids.<br>
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1. Marker<br>
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2-8.EcoRI/PstI digests of pACYC177+OmpA-linker-omega-linker plasmids<br></var>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103019>alpha_linker under PT7 (BBa_K103019)</a></h3>
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<h4>Michał K.</h4>
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<p><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmid from culture inoculated on previous day - <A href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> carrying <a href=http://partsregistry.org/Part:BBa_K103019>alpha_linker under PT7 (BBa_K103019)</a>.</p>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103020>omega_linker under PT7 (BBa_K103020)</a></h3>
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<h4>Michał K.</h4>
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<ol><li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmid from culture inoculated on previous day (<a href=http://partsregistry.org/wiki/index.php?title=Part:pSB2K3>pSB2K3</a> + <a href=http://partsregistry.org/Part:BBa_K103020>omega_linker under PT7 (BBa_K103020)</a>).</li>
 +
<li>Control <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>digest</a> of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found. <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/14_October_2008#fig2">Fig. 2</a>.</li></ol>
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<a name="fig2"><img src="https://static.igem.org/mediawiki/2008/e/ea/Traw_14_10_2008.jpg" width=200></a>
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<var><b>Fig. 2.</b>EcoRI/PstI digests of isolated pSB2K3+omega_linker under PT7 (BBa_K103020) plasmids<br>
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1. Marker<br>
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2-4. Digested plasmids pSB2K3+omega_linker under PT7 (BBa_K103020)<br></var>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103002>AID under pBAD/araC (BBa_K103002)</a></h3>
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<h4>Michał K.</h4>
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<ol><li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> with overnight ligation - <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> + <a href=http://partsregistry.org/Part:BBa_K103002>AID under pBAD/araC (BBa_K103002)</a>.</li>
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<li> Tranformants plating on LB with ampicillin. </li>
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</ol>
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<h3></h3>
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<h3>Preparation of <a href=http://partsregistry.org/Part:BBa_K103017>OmpA_linker_alpha_linker under Plac (BBa_K103017)</a></h3>
<h4>Piotr</h4>
<h4>Piotr</h4>
<ol>
<ol>
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<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> with overnight ligations.</li>
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<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#chemotransform">Transformation</a> of <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">TOP10</a> with overnight ligation - <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> (with OmpA_linker and lactose promoter) + alpha2.</li>
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<li> Tranformants planting on LB with kanamycin - 3kb(?????)+ link_alpha, 3kb(????)_pLac_OmpA_ + alpha2 and RFP(?????) + AraC+pBAD+AID on LB with ampicillin. </li>
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<li> Tranformants plating on LB with kanamycin. </li>
</ol>
</ol>

Latest revision as of 18:57, 28 October 2008

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Preparation of linker_alpha (BBa_K103009)

Piotr

  1. Transformation of TOP10 with overnight ligation - pSB2K3 + linker_alpha (BBa_K103009).
  2. Tranformants plating on LB with kanamycin.

Preparation of OmpA-linker-omega-linker (BBa_K103016)

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day - pACYC177 + OmpA-linker-omega-linker (BBa_K103016).
  2. Control digest of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - still no proper clones found. Fig. 1.
Fig. 1.EcoRI/PstI digests of isolated plasmids.
1. Marker
2-8.EcoRI/PstI digests of pACYC177+OmpA-linker-omega-linker plasmids

Preparation of alpha_linker under PT7 (BBa_K103019)

Michał K.

Isolation of plasmid from culture inoculated on previous day - pSB1A3 carrying alpha_linker under PT7 (BBa_K103019).

Preparation of omega_linker under PT7 (BBa_K103020)

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day (pSB2K3 + omega_linker under PT7 (BBa_K103020)).
  2. Control digest of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found. Fig. 2.
Fig. 2.EcoRI/PstI digests of isolated pSB2K3+omega_linker under PT7 (BBa_K103020) plasmids
1. Marker
2-4. Digested plasmids pSB2K3+omega_linker under PT7 (BBa_K103020)

Preparation of AID under pBAD/araC (BBa_K103002)

Michał K.

  1. Transformation of TOP10 with overnight ligation - pSB1A3 + AID under pBAD/araC (BBa_K103002).
  2. Tranformants plating on LB with ampicillin.

Preparation of OmpA_linker_alpha_linker under Plac (BBa_K103017)

Piotr

  1. Transformation of TOP10 with overnight ligation - pSB2K3 (with OmpA_linker and lactose promoter) + alpha2.
  2. Tranformants plating on LB with kanamycin.

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