Team:Warsaw/Calendar-Main/16 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>Michał K.</h4><ol>
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<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmids from culture inoculated on previous day.</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 with EcoRI and PstI (Orange buffer). Gel electrophoresis -  proper clones found. <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/16_October_2008#fig1">Fig. 1</a>.</li></ol>
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<p class="hide"><img src="https://static.igem.org/mediawiki/2008/f/f7/Traw_16_10_2008.jpg">
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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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2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)<br>
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8-11. Digests of pSB2K3 carrying alpha_linker under pLac<br>
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11-17. Digests of pSB2K3 carrying linker_alpha<br></var></p>
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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>
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<p> Inoculation of colonies from plate with new ligation of <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> to LB with kanamycin. </p>
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<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmids from culture inoculated on previous day 3kb(?????)+ link_alpha, 3kb(????)_pLac_OmpA_ + alpha2 and RFP(?????) + AraC+pBAD+AID.</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: 3kb(?????)+ link_alpha, 3kb(????)_pLac_OmpA_ + alpha2, RFP(?????) + AraC+pBAD+AID and RFP(?????) + pT7_alpha (14 October) with EcoRI and PstI (Orange buffer). Gel electrophoresis - no proper clones found for RFP(?????) + AraC+pBAD+AID ligation but we found good clones for other ligations.</li>
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<li> Inoculation of colonies from plate with ligation of pACYC177 + OmpA_omega_ to LB with kanamycin. </li>
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<li> Repetition of <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of isolated DNA fragments: RFP(?????) + AraC+pBAD+AID.</li>
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<li>Overnight <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> of RFP(?????) + pT7_alpha EcoRI and BcuI (BamHI buffer). </li>
 
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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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<ol><li>Control <a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#digest>digest</a> of isolated plasmid - <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> fragment (<a href=https://2008.igem.org/Team:Warsaw/Calendar-Main/14_October_2008>14 October</a>) with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found. <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/16_October_2008#fig1">Fig. 1</a>.</li>
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<li>Overnight <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> <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> with EcoRI and BcuI (BamHI buffer). </li>
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</ol>
 +
 +
<p class="hide"><img src="https://static.igem.org/mediawiki/2008/f/f7/Traw_16_10_2008.jpg">
 +
<var><b>Fig. 1.</b>EcoRI/PstI digests of isolated plasmids<br>
 +
1. Marker<br>
 +
2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)<br>
 +
8-11. Digests of pSB2K3 carrying alpha_linker under pLac<br>
 +
11-17. Digests of pSB2K3 carrying linker_alpha<br></var></p>
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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#plasmid_DNA_isolation>Isolation</a> of plasmids from culture inoculated on previous day (<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>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/16_October_2008#fig2">Fig. 2</a>.</li>
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<li> Repetition of overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of isolated DNA fragments: <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></ol>
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<a name="fig2"><img src="https://static.igem.org/mediawiki/2008/3/3c/Traw2_15_10_2008.jpg"></a>
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<var><b>Fig. 2.</b>EcoRI/PstI digests of isolated plasmids (pSB1A3+AID under pBAD/araC (BBa_K103002)<br>
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1. Marker<br>
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2-9. EcoRI/PstI digests of isolated plasmids<br></var>
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 +
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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>
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<h4>Michał K.</h4>
 +
<ol>
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<li><a href=https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation>Isolation</a> of plasmids from culture inoculated on previous day - <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + <a href=http://partsregistry.org/Part:BBa_K103017>OmpA_linker_alpha_linker under Plac (BBa_K103017)</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 - we found good clones. <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/16_October_2008#fig1">Fig. 1</a>.</li>
</ol>
</ol>
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<h3></h3>
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<h4>Piotr</h4>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/f/f7/Traw_16_10_2008.jpg" width=300></a>
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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 above ligation.</li>
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<var><b>Fig. 1.</b>EcoRI/PstI digests of isolated plasmids<br>
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<li> Planting on LB with ampicillin.</li></ol>
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1. Marker<br>
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2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)<br>
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8-11. Digests of pSB2K3 carrying alpha_linker under pLac<br>
 +
11-17. Digests of pSB2K3 carrying linker_alpha<br></var>
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 +
 

Latest revision as of 18:58, 28 October 2008

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Preparation of linker_alpha (BBa_K103009)

Michał K.

  1. Isolation of plasmids from culture inoculated on previous day.
  2. Control digest of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found. Fig. 1.

Fig. 1.EcoRI/PstI digests of isolated plasmids
1. Marker
2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)
8-11. Digests of pSB2K3 carrying alpha_linker under pLac
11-17. Digests of pSB2K3 carrying linker_alpha

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

Michał K.

Inoculation of colonies from plate with new ligation of pACYC177 + OmpA-linker-omega-linker (BBa_K103016) to LB with kanamycin.

Preparation of alpha_linker under PT7 (BBa_K103019)

Michał K.

  1. Control digest of isolated plasmid - pSB1A3 carrying alpha_linker under PT7 (BBa_K103019) fragment (14 October) with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found. Fig. 1.
  2. Overnight digest pSB1A3 carrying alpha_linker under PT7 (BBa_K103019) with EcoRI and BcuI (BamHI buffer).

Fig. 1.EcoRI/PstI digests of isolated plasmids
1. Marker
2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)
8-11. Digests of pSB2K3 carrying alpha_linker under pLac
11-17. Digests of pSB2K3 carrying linker_alpha

Preparation of AID under pBAD/araC (BBa_K103002)

Michał K.

  1. Isolation of plasmids from culture inoculated on previous day (pSB1A3 + AID under pBAD/araC (BBa_K103002)).
  2. Control digest of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - still no proper clones found. Fig. 2.
  3. Repetition of overnight ligation of isolated DNA fragments: pSB1A3 + AID under pBAD/araC (BBa_K103002).
Fig. 2.EcoRI/PstI digests of isolated plasmids (pSB1A3+AID under pBAD/araC (BBa_K103002)
1. Marker
2-9. EcoRI/PstI digests of isolated plasmids

Preparation of OmpA_linker_alpha_linker under Plac (BBa_K103017)

Michał K.

  1. Isolation of plasmids from culture inoculated on previous day - pSB2K3 + OmpA_linker_alpha_linker under Plac (BBa_K103017)
  2. Control digest of isolated plasmids with EcoRI and PstI (Orange buffer). Gel electrophoresis - we found good clones. Fig. 1.
Fig. 1.EcoRI/PstI digests of isolated plasmids
1. Marker
2-7. Digests of pSB1A3 carrying alpha_linker under PT7 (BBa_K103019)
8-11. Digests of pSB2K3 carrying alpha_linker under pLac
11-17. Digests of pSB2K3 carrying linker_alpha

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