Team:Goettingen/week14-3

From 2012.igem.org

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<h2><b>V07_30 </b></h2><br>
<h2><b>V07_30 </b></h2><br>
-
<b>2<sup>nd</sup> round: Saturated mutagenesis PCR, 1000 µL</b><br>
+
<b>V07_30_1 2<sup>nd</sup> round: Analysis of the transformation <a href="https://2012.igem.org/Team:Goettingen/week13-3">V07_27</a></b><br>
<ul>
<ul>
-
<li>Experiment: <br>The PCR was set up again, seeing as we lost our DNA material during the ethanol precipitation the previous week.</li>
+
<li>Experiment: <br>The plates and liquid culture were analyzed. </li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>Neither the liquid culture nor the plates exhibited bacterial growth.</li>
 +
</ul>
 +
<br>
 +
<b>V07_30_2 2<sup>nd</sup> round: Saturated mutagenesis PCR, 1000 µL</b><br>
 +
<ul>
 +
<li>Experiment: <br>The PCR was set up again, seeing as we lost our DNA material during the ethanol precipitation the previous week. </li>
</ul>
</ul>
<br></td></tr>
<br></td></tr>
</table>
</table>
 +
<br>
 +
<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
 +
<tr bordercolor="black" valign="top">
 +
<td width="900" bordercolor="black" valign="top">
 +
<h2><b>V07_31 </b></h2><br>
 +
<b>V07_31_1 2<sup>nd</sup> round: PCR clean-up</b><br>
 +
<ul>
 +
<li>Experiment: <br>The PCR clean-up was performed according to the established protocol form <a href="https://2012.igem.org/Team:Goettingen/week10-3">week 10</a>.</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding agarose gel showed a band of the expected size.</li>
 +
</ul>
 +
<br>
 +
<b>V07_31_2 2<sup>nd</sup> round: <i>Dpn</i>I/<i>Bsa</i>I digestion and purification</b><br>
 +
<ul>
 +
<li>Experiment: <br>The digestion was performed with a total volume of 300 µL according to <a href="https://2012.igem.org/Team:Goettingen/week10-3">protocol of week 10</a>. The purification was now again performed using the peqGOLD Cycle-pure Kit (PeqLab). This should stop the loss of DNA material throughout the purification steps!</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding agarose gel showed a band of the expected size.</li>
 +
</ul>
 +
<br>
 +
<b>V07_31_3 2<sup>nd</sup> round: Ligation</b><br>
 +
<ul>
 +
<li>Experiment: <br>The ligation was set up in a 100 µL batch according to <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>. Incubation over night at 16 °C.
 +
</li>
 +
</ul><br>
 +
</td></tr>
 +
</table>
 +
<br>
 +
<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
 +
<tr bordercolor="black" valign="top">
 +
<td width="900" bordercolor="black" valign="top">
 +
<h2><b>V08_01 </b></h2><br>
 +
<b>V08_01_1 2<sup>nd</sup> round: Ethanol precipitation if ligation V07_31</b><br>
 +
<ul>
 +
<li>Experiment: <br>The ethanol precipitation was performed according to the established protocol form <a href="https://2012.igem.org/Team:Goettingen/week10-3">week 10</a>.</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding agarose gel showed a band of the expected size.</li>
 +
</ul>
 +
<br>
 +
<b>V08_01_2 2<sup>nd</sup> round: Transformation of electrocompetent cells with mutant plasmid mixture</b><br>
 +
<ul>
 +
<li>Experiment: <br>Electrocompetent cells were prepared according to <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>. The transformation was performed according to <a href="https://2012.igem.org/Team:Goettingen/week10-3">protocol of week 10</a>.</li>
 +
</ul>
 +
<br>
 +
</td></tr>
 +
</table>
 +
<br>
 +
<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
 +
<tr bordercolor="black" valign="top">
 +
<td width="900" bordercolor="black" valign="top">
 +
<h2><b>V08_02 </b></h2><br>
 +
<b>V08_02_1 2<sup>nd</sup> round: Analysis of transformation V08_01</b><br>
 +
<ul>
 +
<li>Experiment: <br>Plates and liquid culture were checked for successful transformation.</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>No bacterial growth on neither plates nor in liquid culture. Again, our DNA material was presumably lost during the ethanol precipitation. Confirmation see V08__02_2!</li>
 +
</ul>
 +
<br>
 +
<b>V08_02_2 2<sup>nd</sup> round: Test digestion of the ethanol precipitation samples</b><br>
 +
<ul>
 +
<li>Experiment: <br>A test digestion with <i>Eco</i>RI and <i>Pst</i>I was performed on the samples from V08_01_1.</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding agarose gel did not show any bands! There was no DNA material present in the ethanol precipitated samples.</li>
 +
</ul>
 +
<br>
 +
<b>V08_02_3 2<sup>nd</sup> round: mutagenesis PCR from V07_30 repeated</b><br>
 +
<ul>
 +
<li>Experiment: <br>The PCR was performed according to the established protocol from <a href="https://2012.igem.org/Team:Goettingen/week10-3">week 10</a>.</li>
 +
</ul><br>
 +
</td></tr>
 +
</table>
 +
<br>
 +
<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
 +
<tr bordercolor="black" valign="top">
 +
<td width="900" bordercolor="black" valign="top">
 +
<h2><b>V08_03 </b></h2><br>
 +
<b>V08_03_1 2<sup>st</sup> round: PCR clean-up</b><br>
 +
<ul>
 +
<li>Experiment: <br>The clean-up was performed using peqGOLD Gel extraction Kit (PeqLab) following the user manual. 100 µL of elution buffer was used.
 +
</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding gel showed a band of the expected size.
 +
</li>
 +
</ul>
 +
<br>
 +
<b>V08_03_2 2<sup>nd</sup> round: <i>Dpn</i>I/<i>Bsa</i>I digestion</b><br>
 +
<ul>
 +
<li>Experiment: <br>The digestion was performed with a total volume of 200 µL according to <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>.
 +
</li>
 +
</ul><br>
 +
<b>V08_03_3 2<sup>nd</sup> round: Digestion clean-up</b><br>
 +
<ul>
 +
<li>Experiment: <br>The clean-up was performed using peqGOLD Gel extraction Kit (PeqLab) with an EB volume of 100 µL.
 +
</li>
 +
</ul>
 +
<ul>
 +
<li>Observations & Results: <br>The corresponding gel showed a band of the expected size.
 +
</li>
 +
</ul><br></td></tr>
 +
</table>
 +
<br>
 +
<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
 +
<tr bordercolor="black" valign="top">
 +
<td width="900" bordercolor="black" valign="top">
 +
<h2><b>V08_05 </b></h2><br>
 +
<b>2<sup>nd</sup> round: Ligation</b><br>
 +
<ul>
 +
<li>Experiment: <br>The ligation was set up in a 2000 µL batch according to <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>. Incubation over night at 16 °C.
 +
</li>
 +
</ul>
 +
 +
<br></td></tr>
 +
</table>
<br>
<br>
<a href="https://2012.igem.org/Team:Goettingen/Notebook">Back to overview</a><br>
<a href="https://2012.igem.org/Team:Goettingen/Notebook">Back to overview</a><br>
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{{GoettingenFooter}}
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b>V07_31_2 2
 +
b>V07_31_2 2
 +
b>V07_31_2 2
 +
/li>
 +
/li>
 +
b>V07_31_2 2
 +
/li>
 +
b>V08_03_2 2

Latest revision as of 14:52, 26 September 2012

Deutsch  / English 

#3 Chemoreceptor Library - 14th Week

Back to overview

V07_30


V07_30_1 2nd round: Analysis of the transformation V07_27
  • Experiment:
    The plates and liquid culture were analyzed.
  • Observations & Results:
    Neither the liquid culture nor the plates exhibited bacterial growth.

V07_30_2 2nd round: Saturated mutagenesis PCR, 1000 µL
  • Experiment:
    The PCR was set up again, seeing as we lost our DNA material during the ethanol precipitation the previous week.


V07_31


V07_31_1 2nd round: PCR clean-up
  • Experiment:
    The PCR clean-up was performed according to the established protocol form week 10.
  • Observations & Results:
    The corresponding agarose gel showed a band of the expected size.

V07_31_2 2nd round: DpnI/BsaI digestion and purification
  • Experiment:
    The digestion was performed with a total volume of 300 µL according to protocol of week 10. The purification was now again performed using the peqGOLD Cycle-pure Kit (PeqLab). This should stop the loss of DNA material throughout the purification steps!
  • Observations & Results:
    The corresponding agarose gel showed a band of the expected size.

V07_31_3 2nd round: Ligation
  • Experiment:
    The ligation was set up in a 100 µL batch according to protocol. Incubation over night at 16 °C.


V08_01


V08_01_1 2nd round: Ethanol precipitation if ligation V07_31
  • Experiment:
    The ethanol precipitation was performed according to the established protocol form week 10.
  • Observations & Results:
    The corresponding agarose gel showed a band of the expected size.

V08_01_2 2nd round: Transformation of electrocompetent cells with mutant plasmid mixture
  • Experiment:
    Electrocompetent cells were prepared according to protocol. The transformation was performed according to protocol of week 10.


V08_02


V08_02_1 2nd round: Analysis of transformation V08_01
  • Experiment:
    Plates and liquid culture were checked for successful transformation.
  • Observations & Results:
    No bacterial growth on neither plates nor in liquid culture. Again, our DNA material was presumably lost during the ethanol precipitation. Confirmation see V08__02_2!

V08_02_2 2nd round: Test digestion of the ethanol precipitation samples
  • Experiment:
    A test digestion with EcoRI and PstI was performed on the samples from V08_01_1.
  • Observations & Results:
    The corresponding agarose gel did not show any bands! There was no DNA material present in the ethanol precipitated samples.

V08_02_3 2nd round: mutagenesis PCR from V07_30 repeated
  • Experiment:
    The PCR was performed according to the established protocol from week 10.


V08_03


V08_03_1 2st round: PCR clean-up
  • Experiment:
    The clean-up was performed using peqGOLD Gel extraction Kit (PeqLab) following the user manual. 100 µL of elution buffer was used.
  • Observations & Results:
    The corresponding gel showed a band of the expected size.

V08_03_2 2nd round: DpnI/BsaI digestion
  • Experiment:
    The digestion was performed with a total volume of 200 µL according to protocol.

V08_03_3 2nd round: Digestion clean-up
  • Experiment:
    The clean-up was performed using peqGOLD Gel extraction Kit (PeqLab) with an EB volume of 100 µL.
  • Observations & Results:
    The corresponding gel showed a band of the expected size.


V08_05


2nd round: Ligation
  • Experiment:
    The ligation was set up in a 2000 µL batch according to protocol. Incubation over night at 16 °C.


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b>V07_31_2 2 b>V07_31_2 2 b>V07_31_2 2 /li> /li> b>V07_31_2 2 /li> b>V08_03_2 2

Retrieved from "http://2012.igem.org/Team:Goettingen/week14-3"