Team:Warsaw/Calendar-Main/3 October 2008

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<h4>Michał K.</h4>
<h4>Michał K.</h4>
<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:pSB1A3>pSB1A3</a>+<a href=http://partsregistry.org/Part:BBa_K103006>OmpA-linker (BBa_K103006)</a>).</li>
<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:pSB1A3>pSB1A3</a>+<a href=http://partsregistry.org/Part:BBa_K103006>OmpA-linker (BBa_K103006)</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 plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found <li> (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/3_October_2008#fig1>Fig. 1.</a>).</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 plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/3_October_2008#fig1">Fig. 1</a>).</li></ol>
<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/0/0b/Traw_petxba_omp_07_10_2008_na_3_10.jpg"></a>
<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/0/0b/Traw_petxba_omp_07_10_2008_na_3_10.jpg"></a>
<var><b>Fig. 1.</b>Control digest of pSB1A3+OmpA-linker(BBa_K103006)<br>
<var><b>Fig. 1.</b>Control digest of pSB1A3+OmpA-linker(BBa_K103006)<br>
1. Marker<br>
1. Marker<br>
-
2-5. pSB1A3 carrying OmpA-linker (BBa_K103006)</var>
+
2-5. EcoRI/PstI control digest pSB1A3 carrying OmpA-linker (BBa_K103006)</var>
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<ol>  
<ol>  
<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#digest">Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-omega>pET15b+OmpA_omega</a> with XbaI (Tango buffer). DNA ends <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#blunting">blunting</a> with Klenow fragment (3 hr). </li>
<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/protocols#digest">Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-omega>pET15b+OmpA_omega</a> with XbaI (Tango buffer). DNA ends <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#blunting">blunting</a> with Klenow fragment (3 hr). </li>
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<li> Gel electrophoresis (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/3_October_2008#fig2>Fig. 2.</a>) and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper band (<a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-omega>pET15b+OmpA_omega</a> (with removed XbaI site) - 6500 bp).</li>
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<li> Gel electrophoresis and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper band (<a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-omega>pET15b+OmpA_omega</a> (with removed XbaI site) - 6500 bp). <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/3_October_2008#fig2">Fig. 2</a>. </li>
</ol>
</ol>
<a name="fig2"><img src="https://static.igem.org/mediawiki/2008/c/c9/Go_02_10_2008_na_03_10.jpg"></a>
<a name="fig2"><img src="https://static.igem.org/mediawiki/2008/c/c9/Go_02_10_2008_na_03_10.jpg"></a>
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<var><b>Fig. 2.</b>Digest of pET15b+OmpA_omega <br>
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<var><b>Fig. 2.</b>XbaI digest of pET15b+OmpA_omega <br>
1. Marker<br>
1. Marker<br>
-
2. pET15b+OmpA_omega</var>
+
2. XbaI digest pET15b+OmpA_omega</var>

Latest revision as of 20:43, 28 October 2008

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Preparation of linker_alpha (BBa_K103009)

Piotr

  1. Transformation of TOP10 with ligation pSB2K3 + linker_alpha (BBa_K103009).
  2. Plating on LB with kanamycin.

Preparation of linker_omega (BBa_K103013)

Piotr

  1. Transformation of TOP10 with ligation pSB2K3 + linker_omega (BBa_K103013).
  2. Plating on LB with kanamycin.

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

Piotr

  1. Transformation of TOP10 with ligation pACYC177 + OmpA-linker-omega-linker (BBa_K103016).
  2. Plating on LB with kanamycin.

Preparation of OmpA-linker (BBa_K103006)

Michał K.

  1. Isolation of plasmid from culture inoculated on previous day (pSB1A3+OmpA-linker (BBa_K103006)).
  2. Control digest of isolated plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found (Fig. 1).
Fig. 1.Control digest of pSB1A3+OmpA-linker(BBa_K103006)
1. Marker
2-5. EcoRI/PstI control digest pSB1A3 carrying OmpA-linker (BBa_K103006)

Preparation of vector for pT7 constructs

Michał K.

  1. Digest of pET15b+OmpA_omega with XbaI (Tango buffer). DNA ends blunting with Klenow fragment (3 hr).
  2. Gel electrophoresis and gel-out of proper band (pET15b+OmpA_omega (with removed XbaI site) - 6500 bp). Fig. 2.
Fig. 2.XbaI digest of pET15b+OmpA_omega
1. Marker
2. XbaI digest pET15b+OmpA_omega


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