Difference between revisions of "Team:Bielefeld-CeBiTec/Notebook/Labjournal"

 
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> electroporation transformation of BBa_K1416000</p>
 
<p><b>Superior experiment:</b> electroporation transformation of BBa_K1416000</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> electroporation transformation of BBa_KJ36848</p>
 
<p><b>Superior experiment:</b> electroporation transformation of BBa_KJ36848</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_K1416000</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_K1416000</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 86: Line 86:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K1416000</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K1416000</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 103: Line 103:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_J36848</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_J36848</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 120: Line 120:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_J36848</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_J36848</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of BBa_E0400</p>
 
<p><b>Superior experiment:</b> heatshock transformation of BBa_E0400</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_E0400</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_E0400</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_E0400</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_E0400</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of BBa_K525998</p>
 
<p><b>Superior experiment:</b> heatshock transformation of BBa_K525998</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_K525998</p>
 
<p><b>Superior experiment:</b> preculture of transformed BBa_K525998</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K525998</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K525998</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F1,F3,F5</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F1,F3,F5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard gibson protocol</li>
+
<li>  standard <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OS, DK, CDR</p>
+
<p><b>Investigators:</b> Olga Schmidt, Denise Kerkhoff, Christina Drake</p>
 
<p><b>Superior experiment:</b> Overnightculture of barnase</p>
 
<p><b>Superior experiment:</b> Overnightculture of barnase</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and 0,75  µl chloramphenicol</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and 0,75  µl chloramphenicol</li>
 
<li> inkubation overnight 37°C, 140rpm</li>
 
<li> inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of barnase-plasmid</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of barnase-plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR, DK</p>
+
<p><b>Investigators:</b> Christina Drake, Denise Kerkhoff</p>
 
<p><b>Superior experiment:</b> Transformation via Distribution of the parts BBA_J04450, BBa_I746909, BBa_K80800 and BBa_psB1K5</p>
 
<p><b>Superior experiment:</b> Transformation via Distribution of the parts BBA_J04450, BBa_I746909, BBa_K80800 and BBa_psB1K5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  transformation via electroporation</li>
+
<li>  <a target="blank" href="https://static.igem.org/mediawiki/2017/a/ab/T--Bielefeld-CeBiTec--protocol_electrotrafo.pdf">transformation via electroporation</a></li>
 
<li>  plated out: BBa_J04450 on tetracycline, BBa_I74909 and BBa_K808000 on chloramphenicol, BBa_psB1K5 on kanamycin</li>
 
<li>  plated out: BBa_J04450 on tetracycline, BBa_I74909 and BBa_K808000 on chloramphenicol, BBa_psB1K5 on kanamycin</li>
 
</ul>
 
</ul>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR, DK</p>
+
<p><b>Investigators:</b> Christina Drake, Denise Kerkhoff</p>
 
<p><b>Superior experiment:</b> Transformation of BBa_psB6A1</p>
 
<p><b>Superior experiment:</b> Transformation of BBa_psB6A1</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  transformation via electroporation</li>
+
<li>  <a target="blank" href="https://static.igem.org/mediawiki/2017/a/ab/T--Bielefeld-CeBiTec--protocol_electrotrafo.pdf">transformation via electroporation</a></li>
 
<li> plated out on amphenicol</li>
 
<li> plated out on amphenicol</li>
 
</ul>
 
</ul>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> DK, CDR</p>
+
<p><b>Investigators:</b> Denise Kerkhoff, Christina Drake</p>
 
<p><b>Superior experiment:</b> Overnightculture of BBA_J04450, BBa_I746909 and BBa_K808000</p>
 
<p><b>Superior experiment:</b> Overnightculture of BBA_J04450, BBa_I746909 and BBa_K808000</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and at BBA_J04450 3µl tetracycline and atBBa_I746909 and BBa_K808000 0,75  µl chloramphenicol</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and at BBA_J04450 3µl tetracycline and atBBa_I746909 and BBa_K808000 0,75  µl chloramphenicol</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of BBA_J04450, BBa_I746909 and BBa_K80800-plasmids</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of BBA_J04450, BBa_I746909 and BBa_K80800-plasmids</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> DK</p>
+
<p><b>Investigators:</b> Denise Kerkhoff</p>
 
<p><b>Superior experiment:</b> Overnightculture of BBA_psB3K5</p>
 
<p><b>Superior experiment:</b> Overnightculture of BBA_psB3K5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and  2,5 µl kanamycin</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and  2,5 µl kanamycin</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F5,F2,F4</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F5,F2,F4</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F2</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F2</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F4</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F4</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F5</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F2,F4,F5</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F2,F4,F5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard gibson protocol</li>
+
<li>  standard <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 7 clones of BBa_K2201221</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 7 clones of BBa_K2201221</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
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</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 2 of BBa_K2201221</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 2 of BBa_K2201221</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 544: Line 544:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201200 clone 20</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201200 clone 20</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 561: Line 561:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201221 clone 2</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201221 clone 2</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 583: Line 583:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Sequenzingorder barnase</p>
 
<p><b>Superior experiment:</b> Sequenzingorder barnase</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 599: Line 599:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> Construction of pSB1C3-PlacUV5_PtNTT2</p>
 
<p><b>Superior experiment:</b> Construction of pSB1C3-PlacUV5_PtNTT2</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 608: Line 608:
 
<li> Nucleo spin plasmid protocol</li>
 
<li> Nucleo spin plasmid protocol</li>
 
</ul>
 
</ul>
<li> Gibson assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
<li> Transformation via heatshock protocol in <i>E. coli</i> DH5<i>alpha</i></li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a> protocol in <i>E. coli</i> DH5<i>alpha</i></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 619: Line 619:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of the positive selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of the positive selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 648: Line 648:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of T7RNAP from KRX-e.coli</p>
 
<p><b>Superior experiment:</b> PCR of T7RNAP from KRX-e.coli</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Q5 High Fidelity-protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/88/T--Bielefeld-CeBiTec--protocol_PCR_Q5_HF_Polymerase.pdf">Q5 High-Fidelity PCR</a></li>
 
<ul>
 
<ul>
 
<li> annealingtemparatur 64°C, elogationtime 1 min</li>
 
<li> annealingtemparatur 64°C, elogationtime 1 min</li>
 
</ul>
 
</ul>
<li> 1%-agarose-TAE-gel</li>
+
<li> 1%-agarose-<a target="blank" href="https://static.igem.org/mediawiki/2017/d/d4/T--Bielefeld-CeBiTec--protocol_TAE_buffer.pdf">TAE</a>-gel</li>
 
<ul>
 
<ul>
 
<li> 1kb Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye</li>
 
<li> 1kb Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye</li>
Line 669: Line 669:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Plated out Barnase from strain collection</p>
 
<p><b>Superior experiment:</b> Plated out Barnase from strain collection</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> solved DNA in 100 µl SOC-medium</li>
+
<li> solved DNA in 100 µl <a target="blank" href="https://static.igem.org/mediawiki/2017/f/fe/T--Bielefeld-CeBiTec--protocol_SOC.pdf">SOC-medium</a></li>
 
<li> plated out on chloramphenicolplate</li>
 
<li> plated out on chloramphenicolplate</li>
 
<li> overnightincubation at 37°C</li>
 
<li> overnightincubation at 37°C</li>
Line 686: Line 686:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> Construction of pK18mobsacB_codA_del</p>
 
<p><b>Superior experiment:</b> Construction of pK18mobsacB_codA_del</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 705: Line 705:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> General</p>
 
<p><b>Superior experiment:</b> General</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 728: Line 728:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
<li> Transformation via heatshock protocol in <i>E. coli</i> BL21</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a> protocol in <i>E. coli</i> BL21</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 744: Line 744:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 767: Line 767:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 795: Line 795:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of barnase</p>
 
<p><b>Superior experiment:</b> PCR of barnase</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 804: Line 804:
 
<li> annealingtemparatur 58°C, elogationtime 30 s</li>
 
<li> annealingtemparatur 58°C, elogationtime 30 s</li>
 
</ul>
 
</ul>
<li> 1%-agarose-TAE-gel</li>
+
<li> 1%-agarose-<a target="blank" href="https://static.igem.org/mediawiki/2017/d/d4/T--Bielefeld-CeBiTec--protocol_TAE_buffer.pdf">TAE</a>-gel</li>
 
<ul>
 
<ul>
 
<li> 100bp Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye</li>
 
<li> 100bp Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye</li>
Line 822: Line 822:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> NEB Biobrick Assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">BioBrick Assembly</a></li>
 
<ul>
 
<ul>
 
<li> RuBisCoTAG mRFP as upstream part, T7-Terminator as downstream part, pSB1A3 as destination plasmid</li>
 
<li> RuBisCoTAG mRFP as upstream part, T7-Terminator as downstream part, pSB1A3 as destination plasmid</li>
 
</ul>
 
</ul>
<li> Transformation via heatshock and DH5α Cells</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a> and DH5α Cells</li>
 
<ul>
 
<ul>
 
<li> used 5µl from the ligation</li>
 
<li> used 5µl from the ligation</li>
Line 845: Line 845:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Sequenzingorder barnase</p>
 
<p><b>Superior experiment:</b> Sequenzingorder barnase</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 858: Line 858:
 
<li> Cellgrowth with the TyrRS-Heidelberg-plasmid</li>
 
<li> Cellgrowth with the TyrRS-Heidelberg-plasmid</li>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and 0,75  µl chloramphenicol</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and 0,75  µl chloramphenicol</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
Line 869: Line 869:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> transformation via electroporation of cells with T7-Promotor</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/ab/T--Bielefeld-CeBiTec--protocol_electrotrafo.pdf">transformation via electroporation</a> of cells with T7-Promotor</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 884: Line 884:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 901: Line 901:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> used 5µl LB media and kanamycin</li>
+
<li> used 5µl <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB media</a> and kanamycin</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 921: Line 921:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of TyrRS-Heidelberg-plasmid</p>
 
<p><b>Superior experiment:</b> PCR of TyrRS-Heidelberg-plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 927: Line 927:
 
<ul>
 
<ul>
 
<li> plasmidisolation with MN protocol 5</li>
 
<li> plasmidisolation with MN protocol 5</li>
<li> Q5 High Fidelity-protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/88/T--Bielefeld-CeBiTec--protocol_PCR_Q5_HF_Polymerase.pdf">Q5 High-Fidelity PCR</a></li>
 
<ul>
 
<ul>
 
<li> annealingtemparatur 53°C, elogationtime 20s</li>
 
<li> annealingtemparatur 53°C, elogationtime 20s</li>
 
</ul>
 
</ul>
 
<li> Gibbsonassembly with 5µl backbone and 0.5 µl insert</li>
 
<li> Gibbsonassembly with 5µl backbone and 0.5 µl insert</li>
<li> Transformation via heatshoc</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
<li> overnight incubation at 37°C</li>
 
<li> overnight incubation at 37°C</li>
 
</ul>
 
</ul>
Line 943: Line 943:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 960: Line 960:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> GoTaq G2 PCR protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/c/cc/T--Bielefeld-CeBiTec--protocol_colonyPCR_GoTaq_G2.pdf">GoTaq® G2 PCR</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 975: Line 975:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> NEB Biobrick Assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">BioBrick Assembly</a></li>
 
<ul>
 
<ul>
 
<li> digest and ligate RuBisCo parts as upstreampart and T7-Terminator/Promotor as downstreampart and pSB1A3 as destination plasmid</li>
 
<li> digest and ligate RuBisCo parts as upstreampart and T7-Terminator/Promotor as downstreampart and pSB1A3 as destination plasmid</li>
Line 1,004: Line 1,004:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> Interlab study</p>
 
<p><b>Superior experiment:</b> Interlab study</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,014: Line 1,014:
 
<li> 1 µL plasmid from each plate</li>
 
<li> 1 µL plasmid from each plate</li>
 
<ul>
 
<ul>
<li> Transformation via heatshock protocol in <i>E. coli</i> XL1-blue</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a> protocol in <i>E. coli</i> XL1-blue</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,024: Line 1,024:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Transformation via heatshock</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,039: Line 1,039:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,056: Line 1,056:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of T7GFP</p>
 
<p><b>Superior experiment:</b> PCR of T7GFP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Q5 High Fidelity-protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/88/T--Bielefeld-CeBiTec--protocol_PCR_Q5_HF_Polymerase.pdf">Q5 High-Fidelity PCR</a></li>
 
<ul>
 
<ul>
 
<li> Insert: annealingtemparatur 60°C,</li>
 
<li> Insert: annealingtemparatur 60°C,</li>
 
<li> Backbone:elogationtime 30s: annealingtemparatur 60°C, elogationtime 1 min</li>
 
<li> Backbone:elogationtime 30s: annealingtemparatur 60°C, elogationtime 1 min</li>
<li> 1%-agarose-TAE-gel</li>
+
<li> 1%-agarose-<a target="blank" href="https://static.igem.org/mediawiki/2017/d/d4/T--Bielefeld-CeBiTec--protocol_TAE_buffer.pdf">TAE</a>-gel</li>
 
</ul>
 
</ul>
 
<li>  result: PCR didn`t work</li>
 
<li>  result: PCR didn`t work</li>
Line 1,077: Line 1,077:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of T7GFP</p>
 
<p><b>Superior experiment:</b> PCR of T7GFP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 1%-agarose-TAE-gel</li>
+
<li> 1%-agarose-<a target="blank" href="https://static.igem.org/mediawiki/2017/d/d4/T--Bielefeld-CeBiTec--protocol_TAE_buffer.pdf">TAE</a>-gel</li>
 
<li>  result: PCR did work</li>
 
<li>  result: PCR did work</li>
<li> Gibsonassembly</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a></li>
<li> transformation via heatshock</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,095: Line 1,095:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Integration of CDS of BBa_I746909 in pSB1C3</p>
 
<p><b>Superior experiment:</b> Integration of CDS of BBa_I746909 in pSB1C3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,126: Line 1,126:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> Interlab study</p>
 
<p><b>Superior experiment:</b> Interlab study</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,133: Line 1,133:
 
<li> Over-night culture of negative control, positive control, test device 1-6</li>
 
<li> Over-night culture of negative control, positive control, test device 1-6</li>
 
<ul>
 
<ul>
<li> LB-media + Cm were inoculated</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-media</a> + Cm were inoculated</li>
 
<li> Cultures were incubated over night (37°C, 200 rpm)</li>
 
<li> Cultures were incubated over night (37°C, 200 rpm)</li>
 
</ul>
 
</ul>
Line 1,149: Line 1,149:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Integration of CDS of BBa_I746909 in pSB1C3</p>
 
<p><b>Superior experiment:</b> Integration of CDS of BBa_I746909 in pSB1C3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,166: Line 1,166:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> NEB Biobrick Assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">BioBrick Assembly</a></li>
<li> transformation via heatshock</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,182: Line 1,182:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Overnightculture of tRNA-psB1C3</p>
 
<p><b>Superior experiment:</b> Overnightculture of tRNA-psB1C3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and 0,75  µl chloramphenicol</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and 0,75  µl chloramphenicol</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
Line 1,198: Line 1,198:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS Plasmid (without aaRS)</p>
 
<p><b>Superior experiment:</b> aaRS Plasmid (without aaRS)</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Master Mix</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix</li>
 
<ul>
 
<ul>
 
<li> Backbone: 3.28 µL    Ori 1.0: 1.73 µL</li>
 
<li> Backbone: 3.28 µL    Ori 1.0: 1.73 µL</li>
Line 1,220: Line 1,220:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,241: Line 1,241:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> PCR with Q5 Master Mix, Primer jb,jn, 65°C</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/88/T--Bielefeld-CeBiTec--protocol_PCR_Q5_HF_Polymerase.pdf">PCR with Q5 High-Fidelity Polymerase</a> Master Mix, Primer jb,jn, 65°C</li>
 
<li> gelelectrophoresis, 1% Agarose</li>
 
<li> gelelectrophoresis, 1% Agarose</li>
 
<li> band  2000-2500 bp (supposed to be around 2200 bp)</li>
 
<li> band  2000-2500 bp (supposed to be around 2200 bp)</li>
Line 1,261: Line 1,261:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,289: Line 1,289:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> GoTaq PCR protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/c/cc/T--Bielefeld-CeBiTec--protocol_colonyPCR_GoTaq_G2.pdf">GoTaq® G2 PCR</a></li>
 
<li> most have negative results -> create new destination plasmid pSB1A3</li>
 
<li> most have negative results -> create new destination plasmid pSB1A3</li>
 
</ul>
 
</ul>
Line 1,305: Line 1,305:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,315: Line 1,315:
 
<li> bands of pRS31, pRS32, pRS34, pRS36, pRS39 at around 1200-1500 bp</li>
 
<li> bands of pRS31, pRS32, pRS34, pRS36, pRS39 at around 1200-1500 bp</li>
 
</ul>
 
</ul>
<li> GoTaq Master Mix, Primer hq, jk (Gibson primer) with the colonies pRS31, pRS32, pRS34, pRS36, pRS39</li>
+
<li> GoTaq Master Mix, Primer hq, jk (<a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> primer) with the colonies pRS31, pRS32, pRS34, pRS36, pRS39</li>
 
<ul>
 
<ul>
 
<li> pRS31, pRS32, pRS39 : strong bands at around 1200 bp</li>
 
<li> pRS31, pRS32, pRS39 : strong bands at around 1200 bp</li>
Line 1,330: Line 1,330:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,357: Line 1,357:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Aquacloning of tRNA and backbone</p>
 
<p><b>Superior experiment:</b> Aquacloning of tRNA and backbone</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,372: Line 1,372:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,391: Line 1,391:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Master Mix with 5 µl template</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix with 5 µl template</li>
 
<ul>
 
<ul>
 
<li> pSB1C3 fragment (2):      18.5 ng/µL  around  350 bp  2.2 µL</li>
 
<li> pSB1C3 fragment (2):      18.5 ng/µL  around  350 bp  2.2 µL</li>
Line 1,413: Line 1,413:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Master Mix with 5 µL template</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix with 5 µL template</li>
 
<ul>
 
<ul>
 
<li> pSB1C3(nur ori): 22.2  ng/µL  around 2100 bp  1.57 µL</li>
 
<li> pSB1C3(nur ori): 22.2  ng/µL  around 2100 bp  1.57 µL</li>
Line 1,433: Line 1,433:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,450: Line 1,450:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Aquacloning of tRNA and backbone</p>
 
<p><b>Superior experiment:</b> Aquacloning of tRNA and backbone</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,465: Line 1,465:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Gibbsonassembly of barnase and backbone</p>
 
<p><b>Superior experiment:</b> Gibbsonassembly of barnase and backbone</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,480: Line 1,480:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> used 3µl LB media</li>
+
<li> used 3µl <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB media</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,495: Line 1,495:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Overnightculture of tRNA-psB1C3 and barnase-psB1C3</p>
 
<p><b>Superior experiment:</b> Overnightculture of tRNA-psB1C3 and barnase-psB1C3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 3 ml LB-medium and 0,75  µl chloramphenicol</li>
+
<li> 3 ml <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-medium</a> and 0,75  µl chloramphenicol</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
<li>  inkubation overnight 37°C, 140rpm</li>
 
</ul>
 
</ul>
Line 1,511: Line 1,511:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,530: Line 1,530:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of barnase-plasmid and tRNA-plasmids</p>
 
<p><b>Superior experiment:</b> Plasmidisolation of barnase-plasmid and tRNA-plasmids</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,547: Line 1,547:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,573: Line 1,573:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,607: Line 1,607:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Ligation protocol from the NEB Biobrick Assembly protocol</li>
+
<li> Ligation protocol from the <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">BioBrick Assembly</a></li>
 
<ul>
 
<ul>
 
<li> Variation: incubated 1h at RT instead of 10min</li>
 
<li> Variation: incubated 1h at RT instead of 10min</li>
Line 1,624: Line 1,624:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> preparation of KRX-competent cells</p>
 
<p><b>Superior experiment:</b> preparation of KRX-competent cells</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,639: Line 1,639:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CDR</p>
+
<p><b>Investigators:</b> Christina Drake</p>
 
<p><b>Superior experiment:</b> PCR of T7RNA-Polymerase</p>
 
<p><b>Superior experiment:</b> PCR of T7RNA-Polymerase</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> 1%-agarose-TAE-gel</li>
+
<li> 1%-agarose-<a target="blank" href="https://static.igem.org/mediawiki/2017/d/d4/T--Bielefeld-CeBiTec--protocol_TAE_buffer.pdf">TAE</a>-gel</li>
 
<li>  result: PCR did work</li>
 
<li>  result: PCR did work</li>
<li> Gibsonassembly</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a></li>
<li> transformation via heatshock</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,657: Line 1,657:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,680: Line 1,680:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,691: Line 1,691:
 
<li> pRS32:  9.5 ng/µL</li>
 
<li> pRS32:  9.5 ng/µL</li>
 
</ul>
 
</ul>
<li> Gibson Assembly Master Mix with:</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix with:</li>
 
<ul>
 
<ul>
 
<li> 4.1 µL of oK1.1: around 2480 bp  9.5 ng/µL</li>
 
<li> 4.1 µL of oK1.1: around 2480 bp  9.5 ng/µL</li>
Line 1,706: Line 1,706:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F13 and NPA-RS gene synthesis</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F13 and NPA-RS gene synthesis</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,724: Line 1,724:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F13</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F13</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,749: Line 1,749:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F13 and NPA-RS gene synthesis</p>
 
<p><b>Superior experiment:</b> Gibson assembly of F13 and NPA-RS gene synthesis</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard gibson protocol</li>
+
<li>  standard <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,764: Line 1,764:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F5,F1,F3</p>
 
<p><b>Superior experiment:</b> heatshock transformation of assembled fragments F5,F1,F3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,782: Line 1,782:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F1</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F1</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,807: Line 1,807:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F3</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,832: Line 1,832:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> NEB Biobrick Assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">BioBrick Assembly</a></li>
 
<li> repeat digestion of T7-Terminator downstrampart</li>
 
<li> repeat digestion of T7-Terminator downstrampart</li>
 
<li> ligate over night</li>
 
<li> ligate over night</li>
Line 1,849: Line 1,849:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> MED</p>
+
<p><b>Investigators:</b> Maximilian Edich</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Superior experiment:</b> Biobrick Construction with RuBisCo Parts</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> transformation via heatshock</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 1,864: Line 1,864:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F15</p>
 
<p><b>Superior experiment:</b> Q5-PCR, gel and PCR-cleanUp of F15</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,889: Line 1,889:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,911: Line 1,911:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 4 clones of BBa_K2201220</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 4 clones of BBa_K2201220</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,934: Line 1,934:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 3 of BBa_K2201220</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 3 of BBa_K2201220</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,951: Line 1,951:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 1,973: Line 1,973:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest of pSB1C3_I749606</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a> of pSB1C3_I749606</li>
 
<ul>
 
<ul>
 
<li> 5 µL 10x Universal buffer</li>
 
<li> 5 µL 10x Universal buffer</li>
Line 1,994: Line 1,994:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Gibson assembly protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> protocol</li>
<li> Transformation via heatshock protocol in <i>E. coli</i> DH5<i>alpha</i></li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a0/T--Bielefeld-CeBiTec--protocol_heatshock.pdf">transformation via heat shock</a> protocol in <i>E. coli</i> DH5<i>alpha</i></li>
<li> Transformation via electroporation protocol in <i>E. coli</i> DH5<i>alpha</i></li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/ab/T--Bielefeld-CeBiTec--protocol_electrotrafo.pdf">transformation via electroporation</a> protocol in <i>E. coli</i> DH5<i>alpha</i></li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 2,008: Line 2,008:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,031: Line 2,031:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201220 clone 3</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201220 clone 3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,048: Line 2,048:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K608006 and BBa_K2201220</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K608006 and BBa_K2201220</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard biobrick assembly protocol</li>
+
<li>  <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">Standard BioBrick Assembly</a> protocol</li>
 
<ul>
 
<ul>
 
<li>  Backbone: BBa_J04450, 7,7µL</li>
 
<li>  Backbone: BBa_J04450, 7,7µL</li>
Line 2,068: Line 2,068:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of the selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,092: Line 2,092:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product BBa_K2201320</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,110: Line 2,110:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,134: Line 2,134:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 19 clones of BBa_K2201320</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 19 clones of BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,157: Line 2,157:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 19 of BBa_K2201320</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 19 of BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,174: Line 2,174:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K608006 and BBa_K2201221</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K608006 and BBa_K2201221</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard biobrick assembly protocol</li>
+
<li>  <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">Standard BioBrick Assembly</a> protocol</li>
 
<ul>
 
<ul>
 
<li>  Backbone: BBa_J04450, 2,0µL</li>
 
<li>  Backbone: BBa_J04450, 2,0µL</li>
Line 2,194: Line 2,194:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,217: Line 2,217:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,235: Line 2,235:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320 with IPTG</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320 with IPTG</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,254: Line 2,254:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201320 clone 19</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201320 clone 19</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,271: Line 2,271:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,289: Line 2,289:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320 with IPTG</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201320 with IPTG</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,307: Line 2,307:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,330: Line 2,330:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,353: Line 2,353:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product BBa_K2201321</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product BBa_K2201321</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,371: Line 2,371:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Cultivation of BL21(DE3) with BBa_K2201320</p>
 
<p><b>Superior experiment:</b> Cultivation of BL21(DE3) with BBa_K2201320</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  2x 100mL LB-Media with 25µL Cam in 1000mL flasks</li>
+
<li>  2x 100mL <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-media</a> with 25µL Cam in 1000mL flasks</li>
 
<ul>
 
<ul>
 
<li>  innoculation with 3.3 mL preculture of OD 3 to get a starting OD of 0.1</li>
 
<li>  innoculation with 3.3 mL preculture of OD 3 to get a starting OD of 0.1</li>
Line 2,392: Line 2,392:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> tested samples:</li>
 
<li> tested samples:</li>
Line 2,419: Line 2,419:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> tested samples:</li>
 
<li> tested samples:</li>
Line 2,441: Line 2,441:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,458: Line 2,458:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,471: Line 2,471:
 
<li> Elongation: 30 s & 75 s</li>
 
<li> Elongation: 30 s & 75 s</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 5 µL 10x CutSmart buffer</li>
 
<li> 5 µL 10x CutSmart buffer</li>
Line 2,488: Line 2,488:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 3 of BBa_K2201200</p>
 
<p><b>Superior experiment:</b> preculture of the positive clone 3 of BBa_K2201200</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,505: Line 2,505:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> OSC</p>
+
<p><b>Investigators:</b> Olga Schmidt</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Superior experiment:</b> Construction of selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 5 µL NEB2.1 buffer</li>
 
<li> 5 µL NEB2.1 buffer</li>
Line 2,528: Line 2,528:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 32 clones of BBa_K2201200</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 32 clones of BBa_K2201200</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,552: Line 2,552:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> tested samples:</li>
 
<li> tested samples:</li>
Line 2,574: Line 2,574:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201321</p>
 
<p><b>Superior experiment:</b> preculture of BL21(DE3) BBa_K2201321</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,591: Line 2,591:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201321 clone 3</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201321 clone 3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,608: Line 2,608:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> tested samples:</li>
 
<li> tested samples:</li>
Line 2,635: Line 2,635:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system - Primer annealing test</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,641: Line 2,641:
 
<ul>
 
<ul>
 
<li> all samples were diluted to a final concentration of 0.5 µM</li>
 
<li> all samples were diluted to a final concentration of 0.5 µM</li>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 2,651: Line 2,651:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product CP3</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the Biobrick Assembly product CP3</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,669: Line 2,669:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest (<i>Mnl</i>I) of aqua annealing sample: mutC (0.5 µM)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a> (<i>Mnl</i>I) of aqua annealing sample: mutC (0.5 µM)</li>
 
<ul>
 
<ul>
 
<li> 2 µL <i>Mnl</i>I</li>
 
<li> 2 µL <i>Mnl</i>I</li>
Line 2,683: Line 2,683:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> Tested samples:</li>
 
<li> Tested samples:</li>
Line 2,699: Line 2,699:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Screening for positive transformations containing the Biobrick</p>
 
<p><b>Superior experiment:</b> Screening for positive transformations containing the Biobrick</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,722: Line 2,722:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest (<i>Mnl</i>I) of aqua & HEPES annealing sample: mutC (0.5 µM)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a> (<i>Mnl</i>I) of aqua & HEPES annealing sample: mutC (0.5 µM)</li>
 
<ul>
 
<ul>
 
<li> 2 µL <i>Mnl</i>I</li>
 
<li> 2 µL <i>Mnl</i>I</li>
Line 2,736: Line 2,736:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
<ul>
 
<ul>
 
<li> Tested samples:</li>
 
<li> Tested samples:</li>
Line 2,752: Line 2,752:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,779: Line 2,779:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,795: Line 2,795:
 
<li> <i>Eci</i>I -> mutA, oligo2</li>
 
<li> <i>Eci</i>I -> mutA, oligo2</li>
 
</ul>
 
</ul>
<li> Restriction digest:</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a>:</li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 2,804: Line 2,804:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 2,814: Line 2,814:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> SDS-Page and western blot of BBa_K2201321</p>
 
<p><b>Superior experiment:</b> SDS-Page and western blot of BBa_K2201321</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,830: Line 2,830:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,837: Line 2,837:
 
<li> amplification of the mRFP (Phusion Master Mix), Primer vg, vh, annealing temperature: 60°C</li>
 
<li> amplification of the mRFP (Phusion Master Mix), Primer vg, vh, annealing temperature: 60°C</li>
 
<li> amplification of the library backbone(Phusion Master Mix), Primer 17ve, 17vf, Annealing temperature: 61°C</li>
 
<li> amplification of the library backbone(Phusion Master Mix), Primer 17ve, 17vf, Annealing temperature: 61°C</li>
<li> Gibson Assembly with Gibson Master Mix + mRFP (3.1 µL) + library backbone (1.9 µL)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> with Gibson Master Mix + mRFP (3.1 µL) + library backbone (1.9 µL)</li>
 
<li> transformation in DH5a via heatshock</li>
 
<li> transformation in DH5a via heatshock</li>
 
</ul>
 
</ul>
Line 2,848: Line 2,848:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> ssDNA annealing protocoll with the primers 17vi (1 µL), 17vj (1,4 µL)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/d/d1/T--Bielefeld-CeBiTec--protocol_ssDNA_annealing.pdf">ssDNA Annealing</a> protocoll with the primers 17vi (1 µL), 17vj (1,4 µL)</li>
 
<li> agarose gelelectrophoresis, 3 %, positive result: bands of 120 bp</li>
 
<li> agarose gelelectrophoresis, 3 %, positive result: bands of 120 bp</li>
 
</ul>
 
</ul>
Line 2,864: Line 2,864:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle</li>
 
<li> transformation in DH5a via heatshock</li>
 
<li> transformation in DH5a via heatshock</li>
 
</ul>
 
</ul>
Line 2,885: Line 2,885:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 2,906: Line 2,906:
 
<li> <i>Eci</i>I -> mutA, oligo2</li>
 
<li> <i>Eci</i>I -> mutA, oligo2</li>
 
</ul>
 
</ul>
<li> Restriction digest:</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a>:</li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 2,915: Line 2,915:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 2,925: Line 2,925:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle (8 x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle (8 x)</li>
 
<li> transformation in DH5a via heatshock (8 x)</li>
 
<li> transformation in DH5a via heatshock (8 x)</li>
 
</ul>
 
</ul>
Line 2,941: Line 2,941:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (16 x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (16 x)</li>
 
<li> transformation in DH5a via heatshock (16 x)</li>
 
<li> transformation in DH5a via heatshock (16 x)</li>
 
</ul>
 
</ul>
Line 2,957: Line 2,957:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 2,973: Line 2,973:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 2,989: Line 2,989:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 3,005: Line 3,005:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,026: Line 3,026:
 
<li> <i>Eci</i>I -> mutA</li>
 
<li> <i>Eci</i>I -> mutA</li>
 
</ul>
 
</ul>
<li> Restriction digest:</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a>:</li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 3,035: Line 3,035:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 3,050: Line 3,050:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the EutS-part from CU Boulder</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the EutS-part from CU Boulder</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,068: Line 3,068:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the FusionRedTag-part from CU Boulder</p>
 
<p><b>Superior experiment:</b> heatshock transformation of the FusionRedTag-part from CU Boulder</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,086: Line 3,086:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of the EutS-part and FusionRedTag-part from CU Boulder</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of the EutS-part and FusionRedTag-part from CU Boulder</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,104: Line 3,104:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,121: Line 3,121:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,142: Line 3,142:
 
<li> <i>Eci</i>I -> mutA</li>
 
<li> <i>Eci</i>I -> mutA</li>
 
</ul>
 
</ul>
<li> Restriction digest:</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a>:</li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 3,151: Line 3,151:
 
<li> Inactivation: 20 min., 65 °C</li>
 
<li> Inactivation: 20 min., 65 °C</li>
 
</ul>
 
</ul>
<li> Native PAGE standard protocol</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/e/e6/T--Bielefeld-CeBiTec--protocol_native_PAGE.pdf">Native DNA PAGE</a> standard protocol</li>
 
</ul>
 
</ul>
 
</article>
 
</article>
Line 3,161: Line 3,161:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K2201200 in psB3T5</p>
 
<p><b>Superior experiment:</b> Biobrick Assembly of BBa_K2201200 in psB3T5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li>  standard biobrick assembly protocol</li>
+
<li>  <a target="blank" href="https://static.igem.org/mediawiki/2017/8/84/T--Bielefeld-CeBiTec--protocol_Standard_Biobrick.pdf">Standard BioBrick Assembly</a> protocol</li>
 
<ul>
 
<ul>
 
<li>  Backbone: pSB3T5, 2,0µL</li>
 
<li>  Backbone: pSB3T5, 2,0µL</li>
Line 3,184: Line 3,184:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> glycerine stocks of K2201200, K2201220, K2201221, K2201320, K2201321</p>
 
<p><b>Superior experiment:</b> glycerine stocks of K2201200, K2201220, K2201221, K2201320, K2201321</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,198: Line 3,198:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 30 clones of K2201200 in pSB3T5</p>
 
<p><b>Superior experiment:</b> GoTaq PCR of 30 clones of K2201200 in pSB3T5</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,225: Line 3,225:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> Construction of pSB1C3-PlacUV5_PtNTT2</p>
 
<p><b>Superior experiment:</b> Construction of pSB1C3-PlacUV5_PtNTT2</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,254: Line 3,254:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201200 in pSB3T5 and K2201321</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201200 in pSB3T5 and K2201321</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,272: Line 3,272:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201207</p>
 
<p><b>Superior experiment:</b> plasmid isolation of preculture of BBa_K2201207</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,293: Line 3,293:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Superior experiment:</b> M.A.X restriction system</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,316: Line 3,316:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201240 and K2201207</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201240 and K2201207</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,334: Line 3,334:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> YKE</p>
+
<p><b>Investigators:</b> Yannic Kerkhoff</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201241 and K2201207</p>
 
<p><b>Superior experiment:</b> heatshock cotransformation of K2201241 and K2201207</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,361: Line 3,361:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,379: Line 3,379:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 3,411: Line 3,411:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 3,427: Line 3,427:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> positiv selection plasmid</p>
 
<p><b>Superior experiment:</b> positiv selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,445: Line 3,445:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 3,461: Line 3,461:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,507: Line 3,507:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS growth experiment</p>
 
<p><b>Superior experiment:</b> aaRS growth experiment</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,525: Line 3,525:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> positiv selection plasmid</p>
 
<p><b>Superior experiment:</b> positiv selection plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,540: Line 3,540:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 5 µL CutSmart</li>
 
<li> 5 µL CutSmart</li>
Line 3,562: Line 3,562:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,587: Line 3,587:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,615: Line 3,615:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS Plasmid</p>
 
<p><b>Superior experiment:</b> aaRS Plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,623: Line 3,623:
 
<ul>
 
<ul>
 
<li> we stamped out the not randomized, negative, clones, easy to identify by the red colour because of the mRFP</li>
 
<li> we stamped out the not randomized, negative, clones, easy to identify by the red colour because of the mRFP</li>
<li>  washed from the cultivation plates with LB-media</li>
+
<li>  washed from the cultivation plates with <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB-media</a></li>
 
</ul>
 
</ul>
 
<li> restriction of the Library with EcoRI and PstI</li>
 
<li> restriction of the Library with EcoRI and PstI</li>
Line 3,638: Line 3,638:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 3,659: Line 3,659:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,698: Line 3,698:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Superior experiment:</b> aaRS plasmid</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/5/55/T--Bielefeld-CeBiTec--protocol_gibson_assembly.pdf">Gibson Assembly</a> Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
<li> transformation in DH5a via heatshock (25 x)</li>
 
</ul>
 
</ul>
Line 3,714: Line 3,714:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,732: Line 3,732:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 0.5 µL Enzyme</li>
 
<li> 0.5 µL Enzyme</li>
Line 3,775: Line 3,775:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,793: Line 3,793:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 3,823: Line 3,823:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,840: Line 3,840:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
 
<article>
 
<article>
 
<ul>
 
<ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 3 µL Enzyme</li>
 
<li> 3 µL Enzyme</li>
Line 3,873: Line 3,873:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,891: Line 3,891:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,897: Line 3,897:
 
<ul>
 
<ul>
 
<li> Transformation (heatshock) of the library plasmid mix in competent cells containing the positive selction plasmid</li>
 
<li> Transformation (heatshock) of the library plasmid mix in competent cells containing the positive selction plasmid</li>
<li> extended regeneration time: adding 0,5 mM IPTG after 1 h in SOC media</li>
+
<li> extended regeneration time: adding 0,5 mM IPTG after 1 h in <a target="blank" href="https://static.igem.org/mediawiki/2017/f/fe/T--Bielefeld-CeBiTec--protocol_SOC.pdf">SOC media</a></li>
 
<li> using cultivation plates with kan (0.3 mM), Cm (0.25 mM), Tet (0.5 mM), IPTG (0,5 mM)</li>
 
<li> using cultivation plates with kan (0.3 mM), Cm (0.25 mM), Tet (0.5 mM), IPTG (0,5 mM)</li>
 
<ul>
 
<ul>
Line 3,910: Line 3,910:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> LSC</p>
+
<p><b>Investigators:</b> Laura Schlueter</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Superior experiment:</b> pos selection</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,919: Line 3,919:
 
<li> cultivation:</li>
 
<li> cultivation:</li>
 
<ul>
 
<ul>
<li> LB media, Cm (0.25 mM) / Tet (0.5 mM) and 1 mM of the ncAA</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/a/a7/T--Bielefeld-CeBiTec--protocol_LB.pdf">LB media</a>, Cm (0.25 mM) / Tet (0.5 mM) and 1 mM of the ncAA</li>
 
<li> 12 microtiter well plate, 600 rpm, 37°C</li>
 
<li> 12 microtiter well plate, 600 rpm, 37°C</li>
 
</ul>
 
</ul>
Line 3,935: Line 3,935:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 3,953: Line 3,953:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> PCR with Q5-polymerase (per reaction, 25 µL):</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/8/88/T--Bielefeld-CeBiTec--protocol_PCR_Q5_HF_Polymerase.pdf">PCR with Q5 High-Fidelity Polymerase</a>-polymerase (per reaction, 25 µL):</li>
 
<ul>
 
<ul>
 
<li> 5 µL Reaction buffer</li>
 
<li> 5 µL Reaction buffer</li>
Line 3,972: Line 3,972:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 3,998: Line 3,998:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 4,023: Line 4,023:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
 
<li> PCR with innuDRY Standard PCR MasterMix (per reaction, 20 µL):</li>
 
<li> PCR with innuDRY Standard PCR MasterMix (per reaction, 20 µL):</li>
Line 4,040: Line 4,040:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
 
<li> PCR with BioMaster-HS Taq PCR Color (per reaction, 25 µL):</li>
 
<li> PCR with BioMaster-HS Taq PCR Color (per reaction, 25 µL):</li>
Line 4,057: Line 4,057:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
 
<li> PCR with Fire Polymerase (per reaction, 20 µL):</li>
 
<li> PCR with Fire Polymerase (per reaction, 20 µL):</li>
Line 4,074: Line 4,074:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
 
<li> PCR with Phusion polymerase (per reaction, 25 µL):</li>
 
<li> PCR with Phusion polymerase (per reaction, 25 µL):</li>
Line 4,091: Line 4,091:
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<li> PCR and DNA purification kit (Macherey-Nagel)</li>
 
<ul>
 
<ul>
<li> Nucleo spin PCR and DNA Clean-up protocol</li>
+
<li> Nucleo spin PCR and <a target="blank" href="https://static.igem.org/mediawiki/2017/0/0e/T--Bielefeld-CeBiTec--protocol_Monarch_PCR_and_DNA_clean_up_kit_NEB.pdf">Monarch® PCR & DNA Clean-up</a> protocol</li>
 
</ul>
 
</ul>
<li> Restriction digest</li>
+
<li> <a target="blank" href="https://static.igem.org/mediawiki/2017/2/23/T--Bielefeld-CeBiTec--protocol_Restriction_Digest.pdf">Restriction Digest</a></li>
 
<ul>
 
<ul>
 
<li> 1 µL Enzyme</li>
 
<li> 1 µL Enzyme</li>
Line 4,117: Line 4,117:
 
</div>
 
</div>
 
<div class="labnote-content">
 
<div class="labnote-content">
<p><b>Investigators:</b> CMZ</p>
+
<p><b>Investigators:</b> Camilla Maerz</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Superior experiment:</b> PCR with UBP</p>
 
<p><b>Procedure:</b></p>
 
<p><b>Procedure:</b></p>
Line 4,130: Line 4,130:
 
</div>
 
</div>
 
</div>
 
</div>
 +
  
  

Latest revision as of 03:56, 2 November 2017

Labjournal

2017-04-03 - 2017-04-09

getting positive clones for further work

Investigators: Yannic Kerkhoff

Superior experiment: electroporation transformation of BBa_K1416000

Procedure:

  • standard electroporation protocol
    • used electro competend DH5alpha from NEB
    • streaked out on LB-plates with Cam

getting positive clones for further work

Investigators: Yannic Kerkhoff

Superior experiment: electroporation transformation of BBa_KJ36848

Procedure:

  • standard electroporation protocol
    • used electro competend DH5alpha from NEB
    • streaked out on LB-plates with Cam

multiplication of plasmids containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: preculture of transformed BBa_K1416000

Procedure:

  • standard procedure
    • 1,25µl Cam in 5mL LB

2017-04-10 - 2017-04-16

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_K1416000

Procedure:

  • standard protocol
    • 517,9 ng/µL

multiplication of plasmids containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: preculture of transformed BBa_J36848

Procedure:

  • standard procedure
    • 1,25µl Cam in 5mL LB

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_J36848

Procedure:

  • standard protocol
    • 231,8 ng/µL

2017-05-29 - 2017-06-04

getting positive clones for further work

Investigators: Yannic Kerkhoff

Superior experiment: heatshock transformation of BBa_E0400

Procedure:

  • standard heatshock protocol
    • used own produced chemo competend DH5alpha
    • streaked out on LB-plates with Amp

2017-06-05 - 2017-06-11

multiplication of plasmids containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: preculture of transformed BBa_E0400

Procedure:

  • standard procedure
    • 5,0µl Amp in 5mL LB

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_E0400

Procedure:

  • standard protocol
    • 88,4 ng/µL

getting positive clones for further work

Investigators: Yannic Kerkhoff

Superior experiment: heatshock transformation of BBa_K525998

Procedure:

  • standard heatshock protocol
    • used own produced chemo competend DH5alpha
    • streaked out on LB-plates with Cam

multiplication of plasmids containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: preculture of transformed BBa_K525998

Procedure:

  • standard procedure
    • 1,25µl Cam in 5mL LB

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_K525998

Procedure:

  • standard protocol
    • 140,9 ng/µL

2017-06-12 - 2017-06-18

Assemble the fragments to get the Part BBa_K2201220

Investigators: Yannic Kerkhoff

Superior experiment: Gibson assembly of F1,F3,F5

Procedure:

Cellgrowth with the barnaseplasmid

Investigators: Olga Schmidt, Denise Kerkhoff, Christina Drake

Superior experiment: Overnightculture of barnase

Procedure:

  • 3 ml LB-medium and 0,75 µl chloramphenicol
  • inkubation overnight 37°C, 140rpm

To get and purified the plasmid

Investigators: Christina Drake

Superior experiment: Plasmidisolation of barnase-plasmid

Procedure:

  • protocol from Macherley-Nagel: Plasmid DNA purification, protocol 5: NucleoSpin
  • concentration measurement by nanoprop
  • sequenzingorder

Duplicate the parts

Investigators: Christina Drake, Denise Kerkhoff

Superior experiment: Transformation via Distribution of the parts BBA_J04450, BBa_I746909, BBa_K80800 and BBa_psB1K5

Procedure:

Duplicate the part

Investigators: Christina Drake, Denise Kerkhoff

Superior experiment: Transformation of BBa_psB6A1

Procedure:

Cellgrowth with the plasmids

Investigators: Denise Kerkhoff, Christina Drake

Superior experiment: Overnightculture of BBA_J04450, BBa_I746909 and BBa_K808000

Procedure:

  • 3 ml LB-medium and at BBA_J04450 3µl tetracycline and atBBa_I746909 and BBa_K808000 0,75 µl chloramphenicol
  • inkubation overnight 37°C, 140rpm

To get and purified the plasmid

Investigators: Christina Drake

Superior experiment: Plasmidisolation of BBA_J04450, BBa_I746909 and BBa_K80800-plasmids

Procedure:

  • protocol from Macherley-Nagel: Plasmid DNA purification, protocol 5: NucleoSpin
  • concentration measurement by nanoprop:
    • BBA_J04450 26ng/µl and 25,7ng/µl
    • BBa_I746909 90,2ng/µl
    • 104,3 ng/µl -BBa_K808000 118,3 ng/µl
  • and 146,6 ng/µl
  • sequenzingorder

2017-06-19 - 2017-06-25

Cellgrowth with the plasmid

Investigators: Denise Kerkhoff

Superior experiment: Overnightculture of BBA_psB3K5

Procedure:

  • 3 ml LB-medium and 2,5 µl kanamycin
  • inkubation overnight 37°C, 140rpm

getting positive clones for further work

Investigators: Yannic Kerkhoff

Superior experiment: heatshock transformation of assembled fragments F5,F2,F4

Procedure:

  • standard heatshock protocol
    • used own produced chemo competend DH5alpha
    • streaked out on LB-plates with Cam

multiplication, verification and purification of the needed fragment F2: GFPLinker2

Investigators: Yannic Kerkhoff

Superior experiment: Q5-PCR, gel and PCR-cleanUp of F2

Procedure:

  • standard Q5-PCR protocol
    • template: BBa_E0400
    • primer fwd: 17gk
    • primer rev: 17gn
    • annealing temperature: 58°C
    • extension time: 50 seconds
  • standard PCR-cleanUp protocol
    • 125,8 ng/µL

multiplication, verification and purification of the needed fragment F4: StrepLinker2

Investigators: Yannic Kerkhoff

Superior experiment: Q5-PCR, gel and PCR-cleanUp of F4

Procedure:

  • standard Q5-PCR protocol
    • template: BBa_KJ36848
    • primer fwd: 17gp
    • primer rev: 17gq
    • annealing temperature: 61°C
    • extension time: 30 seconds
  • standard PCR-cleanUp protocol
    • 127,0 ng/µL

multiplication, verification and purification of the needed fragment F5: linearized pSB1C3

Investigators: Yannic Kerkhoff

Superior experiment: Q5-PCR, gel and PCR-cleanUp of F5

Procedure:

  • standard Q5-PCR protocol
    • template: K1416000
    • primer fwd: 17gj
    • primer rev: 17gl
    • annealing temperature: 59°C
    • extension time: 120 seconds
  • standard PCR-cleanUp protocol
    • 258,4 ng/µL

Assemble the fragments to get the Part BBa_K2201221

Investigators: Yannic Kerkhoff

Superior experiment: Gibson assembly of F2,F4,F5

Procedure:

Screening for positive transformations containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: GoTaq PCR of 7 clones of BBa_K2201221

Procedure:

  • standard GoTaq-protocol
    • template: colonie pcr clones of GA_K2201221
    • primer fwd: Präfix_fwd
    • primer rev: Suffix_rev
    • annealing temperature: 56°C
    • extension time: 40 seconds
  • positive clone 2

multiplication of plasmids containing the Biobrick

Investigators: Yannic Kerkhoff

Superior experiment: preculture of the positive clone 2 of BBa_K2201221

Procedure:

  • standard procedure
    • 1,25µl Cam in 5mL LB

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_K2201200 clone 20

Procedure:

  • standard protocol
    • 240,6 ng/µL

getting plasmids containing the Biobrick for further experiments

Investigators: Yannic Kerkhoff

Superior experiment: plasmid isolation of preculture of BBa_K2201221 clone 2

Procedure:

  • standard protocol
    • 208,8 ng/µL

2017-06-26 - 2017-07-02

Check the sequence

Investigators: Christina Drake

Superior experiment: Sequenzingorder barnase

Procedure:

  • concentration measurement by nanoprop: 177 ng/µl
  • result: sequence was not barnase

Plasmidisolation and gibson assembly of pSB1C3-PlacUV5_PtNTT2 c30

Investigators: Camilla Maerz

Superior experiment: Construction of pSB1C3-PlacUV5_PtNTT2

Procedure:

Colony PCR of T7RNAP from E. coli KRX

Investigators: Olga Schmidt

Superior experiment: Construction of the positive selection plasmid

Procedure:

  • PCR with Grimson Taq polymerase (per reaction, 25 µL):
    • 5 µL 5x Buffer
    • 2.5 µL dNTPs (2 mM)
    • 0.5 µL 3'-Primer (10 mM)
    • 0.5 µL 5'-Primer (10 mM)
    • 0.125 µL Grimson Taq polymerase
    • 1 picked bacterial colony as template
    • ad 25 µL H20
  • Primer annealing: 64 °C
  • Elongation: 150 s
  • Agarose gel electrophoresis (1%)
    • 100 V, 30 min

Isolate T7RNAP from the chromosome of KRX-e.coli

Investigators: Christina Drake

Superior experiment: PCR of T7RNAP from KRX-e.coli

Procedure:

  • Q5 High-Fidelity PCR
    • annealingtemparatur 64°C, elogationtime 1 min
  • 1%-agarose-TAE-gel
    • 1kb Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye

Plated out Barnase from strain collection

Investigators: Christina Drake

Superior experiment: Plated out Barnase from strain collection

Procedure:

  • solved DNA in 100 µl SOC-medium
  • plated out on chloramphenicolplate
  • overnightincubation at 37°C

Colony PCR of pK18mobsacB-del_codA

Investigators: Camilla Maerz

Superior experiment: Construction of pK18mobsacB_codA_del

Procedure:

  • Go Tag (Promega) protocol
    • Primer: VR, VF2
    • Primer annealing: 56 °C
    • Elongation: 140 s

PCR of pSB1C3

Investigators: Olga Schmidt

Superior experiment: General

Procedure:

  • PCR
    • Q5 High-fidelily (NEB) protocol
    • Denaturation: 64 °C
    • Elongation: 60 s
  • Agarose gel electrophoresis (1%)
    • 100 V, 30 min

Gibson assembly of T7RNAP in pSB1C3

Investigators: Olga Schmidt

Superior experiment: Construction of the selection plasmid

Procedure:

Colony PCR of pSB1C3_T7RNAP c1-8

Investigators: Olga Schmidt

Superior experiment: Construction of the selection plasmid

Procedure:

  • PCR
    • Q5 High-fidelily (NEB) protocol
    • Denaturation: 64 °C
    • Elongation: 80 s
  • Agarose gel electrophoresis (1%)
    • 100 V, 30 min

Colony PCR of barnase (26.06.2017) c1&2

Investigators: Olga Schmidt

Superior experiment: Construction of selection plasmid

Procedure:

  • PCR
    • Go Tag (Promega) protocol
    • Danaturation: 58 °C
    • Elongation: 35 s
  • Agarose gel electrophoresis (1%)
    • 100 V, 30 min

2017-07-03 - 2017-07-09

Check the correct barnasegen

Investigators: Christina Drake

Superior experiment: PCR of barnase

Procedure:

  • Go-Taq-protocol
    • annealingtemparatur 58°C, elogationtime 30 s
  • 1%-agarose-TAE-gel
    • 100bp Marker Geneuler and 5 µl sample with 1 µl 6*Loading Dye
    • result: PCR-product could not be barnase

2017-07-10 - 2017-07-16

Biobrick Assembly of 2B1

Investigators: Maximilian Edich

Superior experiment: Biobrick Construction with RuBisCo Parts

Procedure:

  • BioBrick Assembly
    • RuBisCoTAG mRFP as upstream part, T7-Terminator as downstream part, pSB1A3 as destination plasmid
  • transformation via heat shock and DH5α Cells
    • used 5µl from the ligation
  • plated out on Amp plates and let them grow over night

Check the sequence

Investigators: Christina Drake

Superior experiment: Sequenzingorder barnase

Procedure:

  • result: sequence was not barnase
  • >16.07.2017
  • tRNA
  • Overnightculture of TyrRS-Heidelberg-plasmid
  • CDR
  • Cellgrowth with the TyrRS-Heidelberg-plasmid
    • 3 ml LB-medium and 0,75 µl chloramphenicol
    • inkubation overnight 37°C, 140rpm

    Transformation of T7-Promotor

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    Plasmidisolation of TyrRS-Heidelberg plasmid

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • Plasmid DNA purification kit (Macherey-Nagel)
      • Nucleo spin plasmid protocol

    Preculture of colonies with T7-Promotor

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    2017-07-17 - 2017-07-23

    Isolate the tRNA

    Investigators: Christina Drake

    Superior experiment: PCR of TyrRS-Heidelberg-plasmid

    Procedure:

    Isolation of T7-Promotor

    Investigators: Laura Schlueter

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • Nucleospin Plasmidisolation protocol
      • got four products with the concentrations 50.1 ng/µl, 16.3 ng/µl, 24,2 ng/µl and 2.1 ng/µl

    Colony PCR with 2B1 Colonies

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    Biobrickassembly of 2B2 to 2B8, B1 and A1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • BioBrick Assembly
      • digest and ligate RuBisCo parts as upstreampart and T7-Terminator/Promotor as downstreampart and pSB1A3 as destination plasmid
        • ligate TAG2 and T7-Terminator to 2B2
        • ligate TAG111 and T7-Terminator to 2B3
        • ligate TAG474 and T7-Terminator to 2B4
        • ligate TAG2+TAG111 and T7-Terminator to 2B5
        • ligate TAG2+TAG474 and T7-Terminator to 2B6
        • ligate TAG111+TAG474 and T7-Terminator to 2B7
        • ligate TAG2+TAG111+TAG474 and T7-Terminator to 2B8
        • ligate RuBisCo mRFP and T7-Terminator to B1
        • ligate Carboxysome and T7-Promotor to A1
      • preculture of positive 2B1 colony

    Transformation of test devices

    Investigators: Camilla Maerz

    Superior experiment: Interlab study

    Procedure:

    • dilute test devives (distribution plate 6 & 7) in 10 µL ddH20
    • Transformation

    Transformation of 2B2-2B8, B1 and A1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    Plasmidisolation of 2B1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • Nucleospin Plasmidisolation protocol
      • got 47 ng/µl and 41 ng/µl

    Reverse the insert

    Investigators: Christina Drake

    Superior experiment: PCR of T7GFP

    Procedure:

    • Q5 High-Fidelity PCR
      • Insert: annealingtemparatur 60°C,
      • Backbone:elogationtime 30s: annealingtemparatur 60°C, elogationtime 1 min
      • 1%-agarose-TAE-gel
    • result: PCR didn`t work

    Reverse the insert

    Investigators: Christina Drake

    Superior experiment: PCR of T7GFP

    Procedure:

    PCR of BBa_I746909 and pSB1C3

    Investigators: Olga Schmidt

    Superior experiment: Integration of CDS of BBa_I746909 in pSB1C3

    Procedure:

    • PCR of I746909
      • Q5 High-fidelily (NEB) protocol
      • Primer: 17lk & 17lj
      • Denaturation: 60 °C
      • Elongation: 40 s
    • PCR of pSB1C3
      • Q5 High-fidelily (NEB) protocol
      • Primer: 17ll & 17li
      • Denaturation: 60 °C
      • Elongation: 70 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    Preparation of over-night culture

    Investigators: Camilla Maerz

    Superior experiment: Interlab study

    Procedure:

    • Over-night culture of negative control, positive control, test device 1-6
      • LB-media + Cm were inoculated
      • Cultures were incubated over night (37°C, 200 rpm)

    2017-07-24 - 2017-07-30

    Plasmidisolation of pSB1C3_I746909

    Investigators: Olga Schmidt

    Superior experiment: Integration of CDS of BBa_I746909 in pSB1C3

    Procedure:

    • Plasmid DNA purification kit (Macherey-Nagel)
      • Nucleo spin plasmid protocol

    Repeat the degestion and transformation from the 18th and 19th july

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    Cellgrowth with the plasmid

    Investigators: Christina Drake

    Superior experiment: Overnightculture of tRNA-psB1C3

    Procedure:

    • 3 ml LB-medium and 0,75 µl chloramphenicol
    • inkubation overnight 37°C, 140rpm

    Cloning of the ori (pSB6A1) in pSB1C3

    Investigators: Laura Schlueter

    Superior experiment: aaRS Plasmid (without aaRS)

    Procedure:

    • Gibson Assembly Master Mix
      • Backbone: 3.28 µL Ori 1.0: 1.73 µL
      • Backbone: 0.32 µL Ori 1.1: 4.68 µL
      • Backbone: 0.90 µL Ori 3.0: 4.10 µL
    • Trafo: heatshock
    • not successfull because of the wrong overlapps (try again)

    Colony PCR of the Retrafo of pRS (because of mixed culture)

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Sequencing was not succesfull (maybe mixed colony), so I did a retrafo with the aim of a successful sequencing (incorporation of the aaRS in pSB1C3)
      • pRS 3: 1-5
      • pRS 4: 1-5
      • pRS 5: 1-5
      • pRS 7: 1-5
      • no positive result (no band at 1000 bp)

    Amplification of pSB1C3 for the cloning of the ori in pSB1C3

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • PCR with Q5 High-Fidelity Polymerase Master Mix, Primer jb,jn, 65°C
    • gelelectrophoresis, 1% Agarose
    • band 2000-2500 bp (supposed to be around 2200 bp)
    • Clean up from the gel
      • pSB1C3(nur ori): 22.2 ng/µL

    Amplification of pSB1C3 fragments for a 4 part Gibson (see page 4 in the paper-labbook)

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • PCR, Q5 Master Mix
      • Primer jj, jn for fragment 2, 65°C
      • Primer jb, ht for fragment 4, 65°C
    • gelelectrophoresis, 1% Agarose
      • band at 300 (fragment 2, supposed to be around 326 bp)
      • band at 1200 (fragment 4, supposed to be around 1233 bp)
    • Clean up from the gel
      • fragment 2: 18.5 ng/µL (overlapp(aaRS)_pSB1C3_overlapp(ori))
      • fragment 4: 19.1 ng/µL (overlapp(ori)_pSB1C3_overlapp(aaRS))

    Colony PCR of 2B2-2B8, B1 and A1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • GoTaq® G2 PCR
    • most have negative results -> create new destination plasmid pSB1A3

    Colony PCR of retrafo and more colonies of the pRS

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • GoTaq Master Mix, Primer vr,vf
      • 2 strong bands at 400 bp and 600 bp, small bands of pRS3 6 and pRS4 9 at around 1200-1500 bp
      • bands of pRS31, pRS32, pRS34, pRS36, pRS39 at around 1200-1500 bp
    • GoTaq Master Mix, Primer hq, jk (Gibson Assembly primer) with the colonies pRS31, pRS32, pRS34, pRS36, pRS39
      • pRS31, pRS32, pRS39 : strong bands at around 1200 bp
    • the two positive colony PCRs (vf-vr, hq-jk) for these probes, successful integration of the Tyr-aaRS in pSB1C3 seems to be successfull
    • pRS31, pRS32, pRS39 send to sequencing

    pRS31, pRS32, pRS39 plasmid isolation

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Macherey-Nagel purification kit
      • pRS31: 324.1 ng/µL
      • pRS32: 231.8 ng/µL
      • pRS39: 225.5 ng/µL
    • send tosequencing
    • pRS31, pRS39 have the pSB1C3 with the Tyr-aaRS incorporated (but on position 32, the Arginine (cgt) should be changed to an Leucine (ctg), but it is not. On position 290 there really is a missing Leu, on position 268 the Asparagin is changed to an Arginine)

    2017-07-31 - 2017-08-06

    construct a biobrick

    Investigators: Christina Drake

    Superior experiment: Aquacloning of tRNA and backbone

    Procedure:

    • transformation via heatschock

    plasmid isolation of the positive colonies of the retrafo pRS4 4 (wrong!) and pRS3 12

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Macherey-Nagel purification kit with each 3 mL culture
      • pRS4 4: 44 ng/µL
      • pRS3 12: 68.3 ng/µL
      • not send to sequencing jet

    4 part Gibson and Trafo

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix with 5 µl template
      • pSB1C3 fragment (2): 18.5 ng/µL around 350 bp 2.2 µL
      • pSB1C3 fragment (4): 19.1 ng/µL around 1300 bp 0.8 µL
      • oK1.1 (pMB1 from pSB6A1): 5.8 ng/µL around 1300 bp 1.8 µL
      • Tyr-aaRS: 68.5 ng/µL around 1000 bp 0.2 µL
    • Trafo via heatshock

    Gibson of the ori in pSB1C3 (pori) and Trafo

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix with 5 µL template
      • pSB1C3(nur ori): 22.2 ng/µL around 2100 bp 1.57 µL
      • ok1.0: 5.80 ng/µL around 1258 bp 3.43 µL
    • Trafo via heatshock

    Transfer positive colonies

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • picked and transfered positiv colonies of 2B2, 2B6, 2B7, 2B8 and A1 onto a new plate
    • incubate over night
    • 2B2 and 2B8 are contaminated with negativ colonies

    construct a biobrick

    Investigators: Christina Drake

    Superior experiment: Aquacloning of tRNA and backbone

    Procedure:

    • transformation via heatschock

    construct a biobrick

    Investigators: Christina Drake

    Superior experiment: Gibbsonassembly of barnase and backbone

    Procedure:

    • transformation via heatschock

    Preculture of 2B6, 2B7 and A1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    Cellgrowth with the plasmid

    Investigators: Christina Drake

    Superior experiment: Overnightculture of tRNA-psB1C3 and barnase-psB1C3

    Procedure:

    • 3 ml LB-medium and 0,75 µl chloramphenicol
    • inkubation overnight 37°C, 140rpm

    Plasmidisolation

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • Nucleospin Plasmidisolation protocol
      • got 67.3 ng/µl of 2B6
      • got 32.7 ng/µl of 2B7
      • got 51.6 ng/µl of A1

    To get and purified the plasmid

    Investigators: Christina Drake

    Superior experiment: Plasmidisolation of barnase-plasmid and tRNA-plasmids

    Procedure:

    • protocol from Macherley-Nagel: Plasmid DNA purification, protocol 5: NucleoSpin
    • concentration measurement by nanoprop
    • sequenzingorder

    Redo digestion of the destination plasmid

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • analyze the destination plasmid digestion via gelelectrophoresis
      • pSB1A3 digestion was not effective
    • redo the digestion and analyze it on the gel
      • new dgestion was very effective

    2017-08-07 - 2017-08-13

    Amplification of the backbone (pSB1C3) and insert (Tyr-aaRS)

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • PCR Q5 Polymerase (no Master Mix)
      • for 50 µL reaction
        • 10 µL Reaction Buffer
        • 5 µL 2mM dNTPs
        • 1 µL Template
        • 0,5 µL High Fidelity DNA Polymerase
        • 10.0 µL High GC Enhancer
        • 18.5 µL ddest H2O
        • 5.0 µL Primer
          • pSB1C3 (from RuBisCo, ncAA team): ht, jk 65°C
          • aaRS (from Heidelberg) : hq, jk 65°C (mistake! should be 58°C, redone identically with 58°C)
        • digestion of the PCR products with Dpn1: 5 µL CutSmart Reaktionsbuffer and 1 µL Dpn1 per 50 µL PCR product, on 37°C over night)
      • agarose-gelelectrophhoresis, 1% (bands of the right size)
    • No further use, because results of the sequencing which show that the integration of the incorporation of the Tyr-aaRS in pSB1C3 was already successful (30.06.2017)

    Ligation of 2B2-2B5 and B1

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • Ligation protocol from the BioBrick Assembly
      • Variation: incubated 1h at RT instead of 10min

    characterisation of T7GFP revers

    Investigators: Christina Drake

    Superior experiment: preparation of KRX-competent cells

    Procedure:

    • heatschocktransformation of T7GFP and T7GFP revers in KRX

    construct a biobrick for the selectionplasmid

    Investigators: Christina Drake

    Superior experiment: PCR of T7RNA-Polymerase

    Procedure:

    Amplification of the backbone (pRS31) and insert (ori from pSB6A1, oK1.0) for Gibson assembly

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • PCR Q5 Polymerase (protocoll of 07.08.2017)
      • pRS31: primer jb, jn 65°C
      • oK1: primer jl, jc 65°C
    • digestion with Dpn1 (protocoll of 07.08.2017)
    • agarose-gelelectrophhoresis, 1%
      • band around 1200 bp (oK1.1) / 2500 bp (pRS31), positive

    Gibson Assembly and Trafo of the pRS31 and oK1.0

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • agarose-gelelectrophhoresis, 1%
    • purification from the gel with the Macherey-Nagel purification kit
      • oK1.1: 22.5 ng/µL
      • pRS32: 9.5 ng/µL
    • Gibson Assembly Master Mix with:
      • 4.1 µL of oK1.1: around 2480 bp 9.5 ng/µL
      • 0.9 µL of pRS32: around 1250 bp 22.5 ng/µL
    • Trafo via heatshock

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of assembled fragments F13 and NPA-RS gene synthesis

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend DH5alpha
      • streaked out on LB-plates with Cam

    multiplication, verification and purification of the needed fragment F13: linearized ONBY-Part with removal of the ONBY-RS

    Investigators: Yannic Kerkhoff

    Superior experiment: Q5-PCR, gel and PCR-cleanUp of F13

    Procedure:

    • standard Q5-PCR protocol
      • template: K1416000
      • primer fwd: 17hl
      • primer rev: 17mv
      • annealing temperature: 63°C
      • extension time: 90 seconds
    • standard PCR-cleanUp protocol
      • 80,6 ng/µL

    Assemble the fragments to get the Part BBa_K2201200

    Investigators: Yannic Kerkhoff

    Superior experiment: Gibson assembly of F13 and NPA-RS gene synthesis

    Procedure:

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of assembled fragments F5,F1,F3

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend DH5alpha
      • streaked out on LB-plates with Cam

    multiplication, verification and purification of the needed fragment F1: GFPLinker1

    Investigators: Yannic Kerkhoff

    Superior experiment: Q5-PCR, gel and PCR-cleanUp of F1

    Procedure:

    • standard Q5-PCR protocol
      • template: BBa_E0400
      • primer fwd: 17gk
      • primer rev: 17gm
      • annealing temperature: 58°C
      • extension time: 50 seconds
    • standard PCR-cleanUp protocol
      • 15,9 ng/µL

    multiplication, verification and purification of the needed fragment F3: StrepLinker1

    Investigators: Yannic Kerkhoff

    Superior experiment: Q5-PCR, gel and PCR-cleanUp of F3

    Procedure:

    • standard Q5-PCR protocol
      • template: BBa_KJ36848
      • primer fwd: 17go
      • primer rev: 17gq
      • annealing temperature: 61°C
      • extension time: 30 seconds
    • standard PCR-cleanUp protocol
      • 17,9 ng/µL

    Biobrickassembly

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    • BioBrick Assembly
    • repeat digestion of T7-Terminator downstrampart
    • ligate over night

    Transformation of new plasmids

    Investigators: Maximilian Edich

    Superior experiment: Biobrick Construction with RuBisCo Parts

    Procedure:

    multiplication, verification and purification of the needed fragment F15: linear GLS1

    Investigators: Yannic Kerkhoff

    Superior experiment: Q5-PCR, gel and PCR-cleanUp of F15

    Procedure:

    • standard Q5-PCR protocol
      • template: P2: GLS2
      • primer fwd: 17go
      • primer rev: 17gm
      • annealing temperature: 60°C
      • extension time: 190 seconds
    • standard PCR-cleanUp protocol
      • 36,9 ng/µL

    Primer annealing preparation

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Both Primers (17jh; 17ji) were resuspended in 1x Composite Buffer
    • Dilutions: 100 µM, 10 µM, 5 µM, 2.5 µM, 0.5 µM
    • OD 260 were measured to determine the real concentration

    2017-08-14 - 2017-08-20

    Screening for positive transformations containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: GoTaq PCR of 4 clones of BBa_K2201220

    Procedure:

    • standard GoTaq-protocol
      • template: colonie pcr clones of GA_K2201220
      • primer fwd: Präfix_fwd
      • primer rev: Suffix_rev
      • annealing temperature: 56°C
      • extension time: 40 seconds
    • positive clone 3

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of the positive clone 3 of BBa_K2201220

    Procedure:

    • standard procedure
      • 1,25µl Cam in 5mL LB

    Primer annealing of 17jh & 17ji

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Protocol for annealing oligonucleotides (sigmaaldrich)
      • Eqimolar Primers were mixed (50 µL)
      • Thermal profile (thermocycler)
        • 95 °C, 2 min
        • Cool to 25 °C over 45 min
        • Cool to 4 °C for temporary storage

    Integration of barnase in pSB1C3

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    PCR of T7RNAP_BL

    Investigators: Olga Schmidt

    Superior experiment: Construction of the selection plasmid

    Procedure:

    • PCR
      • Go Tag (Promega) protocol
      • Danaturation: 56 °C
      • Elongation: 180 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    getting plasmids containing the Biobrick for further experiments

    Investigators: Yannic Kerkhoff

    Superior experiment: plasmid isolation of preculture of BBa_K2201220 clone 3

    Procedure:

    • standard protocol
      • 108,9 ng/µL

    Combination of two biobricks to get the composite part BBa_K2201320 for protein expression

    Investigators: Yannic Kerkhoff

    Superior experiment: Biobrick Assembly of BBa_K608006 and BBa_K2201220

    Procedure:

    • Standard BioBrick Assembly protocol
      • Backbone: BBa_J04450, 7,7µL
      • upstream: BBa_K608006, 3,6µL
      • downstram: BBa_K2201220, 5,0µL
      • Ligation time was 12 h at 8 °C

    Colony and plasmid PCR of T7RNAP_BL c1-4

    Investigators: Olga Schmidt

    Superior experiment: Construction of the selection plasmid

    Procedure:

    • PCR
      • Go Tag (Promega) protocol
      • Primer: VR & VF
      • Danaturation: 56 °C
      • Elongation: 180 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of the Biobrick Assembly product BBa_K2201320

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with Cam

    Colony PCR of pSB1C3_barnase (gblock) c1-70

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • PCR
      • Go Tag (Promega) protocol
      • Primer: Vr & VF2
      • Danaturation: 56.5 °C
      • Elongation: 40 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    Screening for positive transformations containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: GoTaq PCR of 19 clones of BBa_K2201320

    Procedure:

    • standard GoTaq-protocol
      • template: colonie pcr clones of GA_K2201320
      • primer fwd: T7-eva_fwd
      • primer rev: Suffix_rev
      • annealing temperature: 51°C
      • extension time: 80 seconds
    • positive clone 19

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of the positive clone 19 of BBa_K2201320

    Procedure:

    • standard procedure
      • 1,25µl Cam in 5mL LB

    Combination of two biobricks to get the composite part BBa_K2201321 for protein expression

    Investigators: Yannic Kerkhoff

    Superior experiment: Biobrick Assembly of BBa_K608006 and BBa_K2201221

    Procedure:

    • Standard BioBrick Assembly protocol
      • Backbone: BBa_J04450, 2,0µL
      • upstream: BBa_K608006, 3,6µL
      • downstram: BBa_K2201221, 2,4µL
      • Ligation time was 12 h at 8°C

    Primer annealing of 17jl & 17jm

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Both Primers were resuspended in ddH20 to a final concentration of 100 µM
    • HEPES annealing standard protocol
    • Aqua annealing standard protocol
      • Final volume: 20 µL
    • Agarose gel electrophoresis (2%)
      • 60 V, 50 min

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of BL21(DE3) BBa_K2201320

    Procedure:

    • standard procedure
      • 1,25µl Cam in 10mL LB
      • 12 h at 37°C

    expression of the fusion protein

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of BL21(DE3) BBa_K2201320 with IPTG

    Procedure:

    • standard procedure
      • 1,25µl Cam in 10mL LB
      • 12 h at 37°C
      • induced with IPTG 0.8mM

    getting plasmids containing the Biobrick for further experiments

    Investigators: Yannic Kerkhoff

    Superior experiment: plasmid isolation of preculture of BBa_K2201320 clone 19

    Procedure:

    • standard protocol
      • 454,4 ng/µL

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of BL21(DE3) BBa_K2201320

    Procedure:

    • standard procedure
      • 1,25µl Cam in 10mL LB
      • 12 h at 37°C

    expression of the fusion protein

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of BL21(DE3) BBa_K2201320 with IPTG

    Procedure:

    • standard procedure
      • 1,25µl Cam in 10mL LB
      • 12 h at 37°C

    Ligation of pSB1C3 and barnase (gblock)

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • Ligation reaction
      • 7 µL vector
      • 3 µL gblock
      • 2 µL T4 DNA ligase buffer
      • 1 µL T4 DNA ligase
      • 17 µL ddH20
    • Incubation: RT, 1 h

    Colony PCR of pSB1C3_barnaseBL c1-60

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • PCR
      • Q5 High-fidelily (NEB) protocol
      • Denaturation: 56.5 °C
      • Elongation: 45 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of the Biobrick Assembly product BBa_K2201321

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with Cam

    - standard cultivation protocoll

    Investigators: Yannic Kerkhoff

    Superior experiment: Cultivation of BL21(DE3) with BBa_K2201320

    Procedure:

    • 2x 100mL LB-media with 25µL Cam in 1000mL flasks
      • innoculation with 3.3 mL preculture of OD 3 to get a starting OD of 0.1
      • incubation for four hours to a OD of 0.6
      • addition of 500µL 0.5mM IPTG each
      • incubation for 8 hours at 37°C and 24 hours at 18°C
      • harveting of the cells

    Native PAGE of annealed primers (17jl & 17jm)

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Native DNA PAGE standard protocol
      • tested samples:
        • HEPES annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
        • Aqua annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
      • Variation: ~40 mA

    2017-08-21 - 2017-08-27

    Native PAGE of annealed primers (17jl & 17jm)

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Native DNA PAGE standard protocol
      • tested samples:
        • HEPES annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
        • Aqua annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
      • Variation: 100 V, 1 h

    Plasmidisolation of pSB1C3_barnaseB c5, 22, 30, 31, 32, 45

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • Plasmid DNA purification kit (Zymo PURE)
      • Nucleo spin plasmid protocol

    PCR of KanR & pSB1C3

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • PCR
      • Q5 High-fidelily (NEB) protocol
      • Primer: 17nx & 17ny (Kan)
      • Primer: 17od & 17oe (pSB1C3)
      • Denaturation: 54 °C & 60 °C
      • Elongation: 30 s & 75 s
    • Restriction Digest
      • 5 µL 10x CutSmart buffer
      • 1 µg plasmid
      • 1 µL DpnI
      • filled up to final volume 50 µL with ddH20
      • Incubation: ON, 37 °C
      • Inactivation: 20 min, 80 °C

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of the positive clone 3 of BBa_K2201200

    Procedure:

    • standard procedure
      • 1,25µl Cam in 5mL LB

    Restriction digest of KanR fragment

    Investigators: Olga Schmidt

    Superior experiment: Construction of selection plasmid

    Procedure:

    • Restriction Digest
      • 5 µL NEB2.1 buffer
      • 2 µL DNA
      • 1 µL XbaI
      • 1 µL PstI
      • filled up to final volume 50 µL with ddH20
      • Incubation: 60 min, 37 °C
      • Heatinactivation: 20 min, 80 °C

    Aim of the Experiment

    Investigators: Yannic Kerkhoff

    Superior experiment: GoTaq PCR of 32 clones of BBa_K2201200

    Procedure:

  • Screening for positive transformations containing the Biobrick
    • standard GoTaq-protocol
      • template: colonie pcr clones of GA_K2201200
      • primer fwd: 17ns
      • primer rev: 17nt
      • annealing temperature: 56°C
      • extension time: 40 seconds
    • positive clone 20

    Native PAGE of annealed primers (17jl & 17jm)

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Native DNA PAGE standard protocol
      • tested samples:
        • HEPES annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
        • Aqua annealing (17jl & 17jm): 100 µM, 10 µM, 5 µM, 2.5 µM, 1 µM, 0.5 µM
      • Variation: 60 V, 2 h

    multiplication of plasmids containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: preculture of BL21(DE3) BBa_K2201321

    Procedure:

    • standard procedure
      • 1,25µl Cam in 5mL LB

    getting plasmids containing the Biobrick for further experiments

    Investigators: Yannic Kerkhoff

    Superior experiment: plasmid isolation of preculture of BBa_K2201321 clone 3

    Procedure:

    • standard protocol
      • 319,1 ng/µL

    Native Page of annealed primers (17jl & 17jm; 17jh & 17ji)

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • Native DNA PAGE standard protocol
      • tested samples:
        • HEPES annealing (17jl & 17jm): 0.5 µM
        • Aqua annealing (17jl & 17jm): 0.5 µM
        • Composite Buffer annealing (17jh & 17 jm): 0.5 µM
        • all Primers (ssDNA): 1 µM

    2017-08-28 - 2017-09-03

    Native PAGE of annealed primers (HEPES annealing & Composite Buffer annealing) of the same concentration to test if all annealings have the same quality

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system - Primer annealing test

    Procedure:

    • all samples were diluted to a final concentration of 0.5 µM
    • Native DNA PAGE standard protocol

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of the Biobrick Assembly product CP3

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with Cam

    Restriction digest of control annealings (mutC)

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Restriction Digest (MnlI) of aqua annealing sample: mutC (0.5 µM)
      • 2 µL MnlI
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 18 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol
      • Tested samples:
        • digested mutC sample
        • native mutC sample
        • ssDNA primers (17jl & 17jm)

    GoTaq PCR of 14 clones of BBa_K2201322

    Investigators: Yannic Kerkhoff

    Superior experiment: Screening for positive transformations containing the Biobrick

    Procedure:

    • standard GoTaq-protocol
      • template: colonie pcr clones of BBA_K2201322
      • primer fwd: Präfix_fwd
      • primer rev: Suffix_rev
      • annealing temperature: 56°C
      • extension time: 40 seconds
    • positive: clone 3, 4, 5

    Restriction digest of aqua & HEPES annealing

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Restriction Digest (MnlI) of aqua & HEPES annealing sample: mutC (0.5 µM)
      • 2 µL MnlI
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 18 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol
      • Tested samples:
        • digested mutC sample (aqua & HEPES annealing)
        • native mutC sample
        • ssDNA primers (17jl & 17jm)

    Aqua annealing of mutA, mutT, mutG and mutC

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Aqua annealing standard protocol (final concentration: 0.5 µM)
      • mutA: 17jf & 17jg
      • mutT: 17jj & 17jk
      • mutG: 17jh & 17ji
      • mutC: 17jl & 17jm
    • final volume of each sample: 50 µL

    2017-09-04 - 2017-09-10

    Restriction digest of control annealings (mutA, mutT, mutG, mutC) and aqua annealing of oligo2

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Oligo2 annealing
      • Aqua annealing standard protocol (final concentration: 0.5 µM)
    • Restricted samples:
      • MnlI -> mutC, oligo2
      • SapI -> mutG, oligo2
      • BsaI -> mutT, oligo2
      • EciI -> mutA, oligo2
    • Restriction Digest:
      • 1 µL Enzyme
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 19 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol

    SDS-Page and western blot of BBa_K2201321

    Investigators: Yannic Kerkhoff

    Superior experiment: SDS-Page and western blot of BBa_K2201321

    Procedure:

    • standard SDS protocoll
    • standard Western blot protocoll

    integration of the mRFP (BBa_J04450) in the aaRS plasmid as optical controle for the integration of an randomerized dsDNA

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • amplification of the mRFP (Phusion Master Mix), Primer vg, vh, annealing temperature: 60°C
    • amplification of the library backbone(Phusion Master Mix), Primer 17ve, 17vf, Annealing temperature: 61°C
    • Gibson Assembly with Gibson Master Mix + mRFP (3.1 µL) + library backbone (1.9 µL)
    • transformation in DH5a via heatshock

    Generating the randomerized dsDNA for the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • ssDNA Annealing protocoll with the primers 17vi (1 µL), 17vj (1,4 µL)
    • agarose gelelectrophoresis, 3 %, positive result: bands of 120 bp

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle
    • transformation in DH5a via heatshock

    2017-09-11 - 2017-09-17

    Annealing, restriction digest and native PAGE of all samples

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Aqua annealing standard protocol (final concentration: 0.5 µM)
      • mutA: 17jf & 17jg
      • mutT: 17jj & 17jk
      • mutG: 17jh & 17ji
      • mutC: 17jl & 17jm
      • oligo2: olgio2_sense & oligo2_antisense
    • final volume of each sample: 50 µL
    • Restricted samples:
      • MnlI -> mutC, oligo2
      • SapI -> mutG, oligo2
      • BsaI -> mutT, oligo2
      • EciI -> mutA, oligo2
    • Restriction Digest:
      • 1 µL Enzyme
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 19 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the optical controle (8 x)
    • transformation in DH5a via heatshock (8 x)

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (16 x)
    • transformation in DH5a via heatshock (16 x)

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    Annealing, restriction digest and native PAGE of all samples

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Aqua annealing standard protocol (final concentration: 0.5 µM)
      • mutA: 17jf & 17jg
      • mutT: 17jj & 17jk
      • mutG: 17jh & 17ji
      • mutC: 17jl & 17jm
      • oligo2: olgio2_sense & oligo2_antisense
    • final volume of each sample: 2x50 µL
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG, oligo2
      • BsaI -> mutT
      • EciI -> mutA
    • Restriction Digest:
      • 1 µL Enzyme
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 19 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol

    2017-09-18 - 2017-09-24

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of the EutS-part from CU Boulder

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend DH5alpha
      • streaked out on LB-plates with Amp

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock transformation of the FusionRedTag-part from CU Boulder

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend DH5alpha
      • streaked out on LB-plates with Kan

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock cotransformation of the EutS-part and FusionRedTag-part from CU Boulder

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend DH5alpha
      • streaked out on LB-plates with AmpKan

    Agarose gel electrophoresis of digested mutA, mutT, mutG & mutC

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Agarose gel electrophoresis (2%)
      • 60 V, 40 min

    Annealing, restriction digest and native PAGE of all samples

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Aqua annealing standard protocol (final concentration: 0.5 µM)
      • mutA: 17jf & 17jg
      • mutT: 17jj & 17jk
      • mutG: 17jh & 17ji
      • mutC: 17jl & 17jm
      • oligo2: olgio2_sense & oligo2_antisense
    • final volume of each sample: 2x50 µL
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG, oligo2
      • BsaI -> mutT
      • EciI -> mutA
    • Restriction Digest:
      • 1 µL Enzyme
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 19 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Native DNA PAGE standard protocol

    Combination of two biobricks to get the part BBa_K2201200 in a tetracycline low copy plasmid

    Investigators: Yannic Kerkhoff

    Superior experiment: Biobrick Assembly of BBa_K2201200 in psB3T5

    Procedure:

    • Standard BioBrick Assembly protocol
      • Backbone: pSB3T5, 2,0µL
      • upstream: BBa_K2201200, 3,6µL
      • Ligation time was 12 h at 8°C
      • cultivation

    glycerine stocks of K2201200, K2201220, K2201221, K2201320, K2201321

    Investigators: Yannic Kerkhoff

    Superior experiment: glycerine stocks of K2201200, K2201220, K2201221, K2201320, K2201321

    Procedure:

  • glycerine stock standard protocol
  • Screening for positive transformations containing the Biobrick

    Investigators: Yannic Kerkhoff

    Superior experiment: GoTaq PCR of 30 clones of K2201200 in pSB3T5

    Procedure:

    • standard GoTaq-protocol
      • template: colonie pcr clones of K2201200 in pSB3T5
      • primer fwd: Präfix_fwd
      • primer rev: Suffix_rev
      • annealing temperature: 56°C
      • extension time: 40 seconds
    • positive clones 1, 3, 6
    • Preculture

    Colony PCR of pSB3K5_mRFP_link_ccdB and pSB1C3_PtNTT2(31-575)-GFP

    Investigators: Camilla Maerz

    Superior experiment: Construction of pSB1C3-PlacUV5_PtNTT2

    Procedure:

    • PCR
      • Go Tag (Promega) protocol
      • Primer: VR, VF2
      • Danaturation: 56 °C
      • Elongation: 90 s & 180 s
    • Agarose gel electrophoresis (1%)
      • 100 V, 30 min

    2017-09-25 - 2017-10-01

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock cotransformation of K2201200 in pSB3T5 and K2201321

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with TetCam

    getting plasmids containing the Biobrick for further experiments

    Investigators: Yannic Kerkhoff

    Superior experiment: plasmid isolation of preculture of BBa_K2201207

    Procedure:

    • standard protocol
      • 75,6 ng/µL
      • Biobrick Assembly

    Aqua annealing of mutA-1, mutG-1, mutC-1, mutG-1 & oligo1

    Investigators: Camilla Maerz

    Superior experiment: M.A.X restriction system

    Procedure:

    • Aqua annealing standard protocol (final concentration: 0.5 µM)
      • mutA-1: 17iv & 17iw
      • mutT-1: 17jb & 17jc
      • mutG-1: 17ix & 17iy
      • mutC-1: 17jd & 17jc
      • oligo1: olgio1_sense & oligo1_antisense
    • final volume: 50 µL

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock cotransformation of K2201240 and K2201207

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with AmpCam

    getting positive clones for further work

    Investigators: Yannic Kerkhoff

    Superior experiment: heatshock cotransformation of K2201241 and K2201207

    Procedure:

    • standard heatshock protocol
      • used own produced chemo competend BL21(DE3)
      • streaked out on LB-plates with AmpCam
      • Q5-PRC + Gel + cleanUp

    2017-10-02 - 2017-10-08

    UBP-PCR with TiTaq

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG, mutC and oligo 2 (1 ng/µL)
      • Primer: 17vt & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • Restriction Digest
      • 1 µL Enzyme
      • 5 µL CutSmart
      • 25 µL mutC dsDNA
      • 19 µL nuclease-free H2O
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG
      • BsaI -> mutT
      • EciI -> mutA
      • EciI, BsaI, SapI, MnlI -> oligo2
      • Variation: oligo2 was digested with 0.5 µL of each enzyme
    • Agarose gel electrophoresis (2 %)
      • 100 V, 20 min

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    Kanymycin with amber stop codon

    Investigators: Laura Schlueter

    Superior experiment: positiv selection plasmid

    Procedure:

    • amplification (~850 bp) of the kanamycin with primers 17jo, 17jp (no second MET codon) 71°C
    • amplification (~850 bp) of the kanamycin with primers 17jr, 17jp (two amber stop codon) 71°C
  • amplification of the K608007 backbone with the primers 17hb, 17hd 57°C
  • Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    UBP-PCR with GoTaq G2 and TiTaq

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG, mutC and oligo 2 (5 ng/µL)
      • Primer: 17 vt & 17 vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG and mutC (5 ng/µL)
      • Primer: VR & VF2
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR with GoTaq:
      • PCR-UBP standard protocol (GoTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG, mutC and oligo2 (5 ng/µL)
      • Primer: 17 vt & 17 vu
      • Primer annealing: 56 °C
      • Elongation: 20 s

    cloning of the aaRS in pSB3C5 (low copy)

    Investigators: Laura Schlueter

    Superior experiment: aaRS growth experiment

    Procedure:

    • restriction of pSB3C5 and aaRS+tRNA (NPA, AcF, Prk) with EcoRI and PstI following the restriction protocol
    • extraction from the gel (Macherey-Nagel Kit)
    • ligation of pSB3C5 and aaRS+tRNA (NPA, AcF, Prk) with T4 Ligase (NEB)
    • Trafo via heatshock

    sceening of the kanamycin (03.10.2017)

    Investigators: Laura Schlueter

    Superior experiment: positiv selection plasmid

    Procedure:

    • Colony PCR (GoTaq, Primer VF2, VR, 56.5 °C)

    Restriction digest of ligated oligo2

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • Restriction Digest
      • 5 µL CutSmart
      • 30 µL ligated oligo2 DNA
      • 2 µL EcoRV
      • filled up to final volume 50 µL with ddH20
      • Incubation: 120 min, 37 °C
      • Inactivation: 20 min, 65 °C

    Gradient PCR of mutT

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR
    • PCR-UBP standard protocol (GoTaq G2)
      • sample: mutT (5 ng/µL)
      • Primer: 17 vt & 17 vu
      • Primer annealing: 50 - 56 °C
      • Elongation: 20 s
    • Agarose gel electrophoresis (2%)
      • 100 V, 30 min

    PCR of mutA, mutT, mutG & mutC with UBP and oligo2 without UBP

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG and mutC (5 ng/µL), oligo2 (25 ng/µL)
      • Primer: VR & VF2
      • Primer annealing: 56 °C
      • Elongation: 20 s
      • Variation:
        • oligo2 without UBP, instead 1 µL H20 was added
        • 0.5 µL isoG & 0.5 µL isoCMe were added to mutA, mutT, mutG and mutC

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS Plasmid

    Procedure:

    • plasmid isolation of the Tyr-RS library-mix
      • we stamped out the not randomized, negative, clones, easy to identify by the red colour because of the mRFP
      • washed from the cultivation plates with LB-media
    • restriction of the Library with EcoRI and PstI
    • proof of the restriction via agarose-gelelectrophoresis, 1 %
      • positve result: bands on ~2000 bp (backbone) and ~1000 bp (Tyr-RS with NNK)

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    2017-10-09 - 2017-10-15

    PCR with switched primers: VF2 & 17vu, VR & 17vt

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG and mutC (5 ng/µL), oligo2 (25 ng/µL)
      • Primer: VF2 & 17vu, VR & 17vt
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR with GoTaq:
      • PCR-UBP standard protocol (GoTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG and mutC (5 ng/µL), oligo2 (25 ng/µL)
      • Primer: VF2 & 17vu, VR & 17vt
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • Agarose gel electrophoresis (2%)
      • 100 V, 30 min

    Generating the library

    Investigators: Laura Schlueter

    Superior experiment: aaRS plasmid

    Procedure:

    • Gibson Assembly Master Mix + dsDNA (3.5 µl) + library Plasmid (0.5 µL) with the opotical controle (25x)
    • transformation in DH5a via heatshock (25 x)

    PCR with higher DNA template concentrations and gradient PCR of mutT

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutT (5 ng/µL) and oligo2 (50 ng/µL)
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • Restriction Digest
      • 0.5 µL Enzyme
      • 2.3 µL CutSmart
      • 20 µL dsDNA
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Restricted samples:
      • BsaI -> mutT
      • EciI, BsaI, SapI, MnlI -> oligo2
      • Variation: oligo2 was digested with 0.5 µL of each enzyme
    • Agarose gel electrophoresis (2 %)
      • 100 V, 20 min
    • Gradient PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutT (5 ng/µL)
      • Primer: VF2 & 17vu
      • Primer annealing: 50 - 58 °C
      • Elongation: 20 s
    • Agarose gel electrophoresis (2%)
      • 100 V, 30 min

    PCR of mutA, mutT, mutG, mutC and oligo2

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with GoTaq:
      • PCR-UBP standard protocol (GoTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG, mutC (5 ng/µL) and oligo2 (25 ng/µL)
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • Restriction Digest
      • 1 µL Enzyme
      • 2.3 µL CutSmart
      • 20 µL dsDNA
      • Incubation: 37 °C, 12 h
      • Inactivation: 20 min., 65 °C
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG
      • BsaI -> mutT
      • EciI -> mutA
      • EciI, BsaI, SapI, MnlI -> oligo2
    • Agarose gel electrophoresis (2 %)
      • 100 V, 30 min

    positive selection with the library

    Investigators: Laura Schlueter

    Superior experiment: pos selection

    Procedure:

    • using cultivation plates with kan (0.5 mM), Cm (0.25 mM), Tet (0.5 mM)
      • no growth

    Variation of restriction digest

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • Restriction Digest
      • 3 µL Enzyme
      • 2.5 µL CutSmart
      • 20 µL dsDNA
      • Incubation: 37 °C, 1 h
      • Inactivation: 20 min., 65 °C
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG
      • BsaI -> mutT
      • EciI -> mutA
      • EciI, BsaI, SapI, MnlI -> oligo2
    • Agarose gel electrophoresis (2 %)
      • 100 V, 30 min

    positive selection with the library

    Investigators: Laura Schlueter

    Superior experiment: pos selection

    Procedure:

    • Transformation (heatshock) of the library plasmid mix in competent cells containing the positive selction plasmid
    • using cultivation plates with kan (0.3 mM), Cm (0.25 mM), Tet (0.5 mM)
      • no growth

    positive selection with the library

    Investigators: Laura Schlueter

    Superior experiment: pos selection

    Procedure:

    • Transformation (heatshock) of the library plasmid mix in competent cells containing the positive selction plasmid
    • extended regeneration time: adding 0,5 mM IPTG after 1 h in SOC media
    • using cultivation plates with kan (0.3 mM), Cm (0.25 mM), Tet (0.5 mM), IPTG (0,5 mM)
      • no growth

    growth experiments with the aaRS

    Investigators: Laura Schlueter

    Superior experiment: pos selection

    Procedure:

    • comparing AcF-TAG, AcF-Leu, Cou-AS, Prk, NPA (on low and high copy plasmid) pSB1C3, pSB3T5
      • cultivation:
        • LB media, Cm (0.25 mM) / Tet (0.5 mM) and 1 mM of the ncAA
        • 12 microtiter well plate, 600 rpm, 37°C

    2017-10-16 - 2017-10-22

    PCR with TiTaq and Q5-polymerase

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • PCR with TiTaq:
      • PCR-UBP standard protocol (TiTaq)
      • samples:
        • Ligation of: mutA, mutT, mutG & mutC (5 ng/µL) and oligo2 (50 ng/µL)
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • PCR with Q5 High-Fidelity Polymerase-polymerase (per reaction, 25 µL):
      • 5 µL Reaction buffer
      • 1.25 µL dNTPs (2 mM)
      • 1.25 µL 3'-Primer (10 mM)
      • 1.25 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 0.25 µL Q5 Hifi-Polymerase
      • 14 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 10 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • Restriction Digest
      • 1 µL Enzyme
      • 20 µL dsDNA
      • 2.3 µL CutSmart Buffer
      • Incubation: 37 °C, 15 h (BsaI & MnlI), 2 h (SapI & EciI)
      • Inactivation: 65 °C, 20 min
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG
      • BsaI -> mutT
      • EciI -> mutA
      • EciI, BsaI, SapI, MnlI -> oligo2

    PCR with different polymerases

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • samples:
      • Ligation of: mutA, mutT, mutG & mutC (5 ng/µL) and oligo2 (50 ng/µL)
    • PCR with Allin Hifi DNA Polymerase (per reaction, 25 µL):
      • 5 µL ReactionBuffer
      • 0.25 µL Hifi DNA Polymerase
      • 2.5 µL 3'-Primer (10 mM)
      • 2.5 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 12.75 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 10 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • PCR with innuDRY Standard PCR MasterMix (per reaction, 20 µL):
      • 10 µL innduDRY Mastermix
      • 2 µL 3'-Primer (10 mM)
      • 2 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 5 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 54 °C
      • Elongation: 30 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • PCR with BioMaster-HS Taq PCR Color (per reaction, 25 µL):
      • 12.5 µL Mastermix
      • 0.7 µL 3'-Primer (10 mM)
      • 0.7 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 10.1 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 51 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • PCR with Fire Polymerase (per reaction, 20 µL):
      • 4 µL Mastermix
      • 0.5 µL 3'-Primer (10 mM)
      • 0.5 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 14 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • PCR with Phusion polymerase (per reaction, 25 µL):
      • 12.5 µL Mastermix
      • 1.25 µL 3'-Primer (10 mM)
      • 1.25 µL 5'-Primer (10 mM)
      • 1 µL DNA template
      • 9 µL H20
      • mutA, mutT, mutG, mutC: + 1 µL H20
      • oligo2: + 0.5 µL isoCme, + 0.5 µL isoG
      • Primer: VF2 & 17vu
      • Primer annealing: 56 °C
      • Elongation: 20 s
    • PCR and DNA purification kit (Macherey-Nagel)
    • Restriction Digest
      • 1 µL Enzyme
      • 20 µL dsDNA
      • 2.3 µL CutSmart Buffer
      • Incubation: 37 °C, 15 h (BsaI & MnlI), 2 h (SapI & EciI)
      • Inactivation: 65 °C, 20 min
    • Restricted samples:
      • MnlI -> mutC
      • SapI -> mutG
      • BsaI -> mutT
      • EciI -> mutA
      • EciI, BsaI, SapI, MnlI -> oligo2

    PCR with different polymerases

    Investigators: Camilla Maerz

    Superior experiment: PCR with UBP

    Procedure:

    • Agarose gel electrophoresis with all samples (17.10)
    • Agarose gel electrophoresis (2%)
      • 100 V, 40 min