Team:Braunschweig/Notebook

From 2013.igem.org

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Yesterday's ligations transformed into <i>E. coli</i> XL1 MRF' by heatshock and plated on 2xYT agar containing glucose and Chloramphenicol.<br>  
Yesterday's ligations transformed into <i>E. coli</i> XL1 MRF' by heatshock and plated on 2xYT agar containing glucose and Chloramphenicol.<br>  
Since the chromoprotein DNA from Uppsala iGEM Team 2011 arrived today we now switched to our new cloning strategy starting with resuspending and transforming the newly arrived DNA into <i>E. coli</i> XL1 Blue MFR' by heatshock. Cells were as well plated on 2xYT agar containing glucose and chloramphenicol.<p>
Since the chromoprotein DNA from Uppsala iGEM Team 2011 arrived today we now switched to our new cloning strategy starting with resuspending and transforming the newly arrived DNA into <i>E. coli</i> XL1 Blue MFR' by heatshock. Cells were as well plated on 2xYT agar containing glucose and chloramphenicol.<p>
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<p><img alt="Chromoprotein arrival" src="https://static.igem.org/mediawiki/2013/2/2e/Braunschweig_Lab_Week7_Chromoproteina_arrival.jpg" height="140px" align="left" vspace="0" hspace="10" style="margin-right:5px"/>
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<p><img alt="Chromoprotein arrival" src="https://static.igem.org/mediawiki/2013/2/2e/Braunschweig_Lab_Week7_Chromoproteina_arrival.jpg" height="150px" align="left" vspace="0" hspace="10" style="margin-right:5px"/>
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Even if we switched to our new strategy we still kept working on the fluorescence markers. Therefore we digested the BioBricks E0420 (eCFP), E0430 (YFP) and J06702 (mCherry) as well as our promotor-RBS-LasR and promotor-RBS-RhlR-constructs. Still lacking the construct containing LuxR we decided to proceed with the promotor-RBS-construct instead to ligate it to the YFP-Brick. Gel extraction was performed for the inserts and DNA purification for the vector parts. Inserts and bricks were ligated overnight using T4 DNA ligase (NEB) at 16°C.<br><br>
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Even if we switched to our new strategy we still kept working on the fluorescence markers. Therefore we digested the BioBricks E0420 (eCFP), E0430 (YFP) and J06702 (mCherry) as well as our promotor-RBS-LasR and promotor-RBS-RhlR-constructs. Still lacking the construct containing LuxR we decided to proceed with the promotor-RBS-construct instead to ligate it to the YFP-Brick. Gel extraction was performed for the inserts and DNA purification for the vector parts. Inserts and bricks were ligated overnight using T4 DNA ligase (NEB) at 16°C.<br><br><br>
Since we now have our first constructs containing the inducible promotors as well as the the ampicillin resistence gen we started our first leakiness experiments. All constructs (P<sub>las</sub>, P<sub>rhl</sub>, P<sub>lux</sub> in combination with the RBS and <i>ampR</i>) were cultivated overnight in 2xYT medium containing various ampicillin concentrations.</p>
Since we now have our first constructs containing the inducible promotors as well as the the ampicillin resistence gen we started our first leakiness experiments. All constructs (P<sub>las</sub>, P<sub>rhl</sub>, P<sub>lux</sub> in combination with the RBS and <i>ampR</i>) were cultivated overnight in 2xYT medium containing various ampicillin concentrations.</p>

Revision as of 23:01, 4 October 2013

Labjournal

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Braunschweig Labbook This is the documentation of our lab work. Achievements of each week are summerized followed by a daily discription of our experiments.

An overview on how we approached this project can be found under Approach. For detailed protocols of certain procedures please refer to Protocols. Attributions are given for each day, however please check our
Attributions section for efforts beyond the lab work.


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