Difference between revisions of "Team:Munich/thisisatest.html"

Revision as of 10:31, 17 October 2018

Transforming E.Coli NEB Turbo to amplify plasmids for pRED/ET Engineering

2018/05/07

Participants:	Dominic Schwarz
Protocol:	Electro-transformation
Notes:	pkD3 contains resistance cassettes with FRT-sites for pRED engineering. Due to the temperature sensitive promotor pNPTS138-R6KT which is used for bone-knockin via RecA recombineering, the cells were incubated at 30°C overnight.
Results:	No colonies. We suspected electroporation to be a problem and switched to chemical transformation in NEB Turbo next.

Redo: Transforming E.Coli NEB Turbo to amplify plasmids for pRED/ET Engineering

2018/05/18

Participants:	Dominic Schwarz
Protocol:	Chemical Transformation
Notes:	Inoculate pRED at 30°C because of temperature sensitive promoter
Results:	No colonies. We suspected the E. Coli NEB Turbo to be a problem and switched to E. Coli Dh5α.

Redo: Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering

2018/05/24

Participants:	Dominic Schwarz
Protocol:	Electro-transformation
Notes:	inoculate pRED at 30°C because of temperature sensitive promoter, pKD3 contains resistance cassette flanked by FRT sites
Results:	We got cells containing the plasmid from PD Dr. Jürgen Lassak of the LMU.

Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering

2018/05/25

Participants:	Dominic Schwarz
Protocol:	Chemical transformation
Notes:	pKD3 contains resistance cassette flanked by FRT sites
Results:	No colonies. Because pRED/ET could not be transformed, we got readily transformed cells from PD Dr. Jürgen Lassak from the LMU

DNA preparation for pRED/ET Engineering

2018/05/26

Participants:	Dominic Schwarz
Protocol:	Miniprep
Notes:	The cells from PD Dr. Jürgen Lassak were used for DNA preparation.
Results:	Concentration of the plasmids obtains from MIniprep: pRED/ET: 37,5 ng/µl pNPTS138-R6KT: 60ng/µl

@@ Line 13: / Line 13: @@
        <td>Protocol:</td>
        <td>
-<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electrocompetent transformation</a>
+<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electro-transformation</a>
 </td>
      </tr>
      <tr>
        <td>Notes:</td>
-       <td>pkD3 contains resistance cassettes with FRT-sites for pRED engineering <br>
+       <td>pkD3 contains resistance cassettes with FRT-sites for pRED engineering. Due to the temperature sensitive promotor pNPTS138-R6KT which is used for bone-knockin via RecA recombineering, the cells were incubated at 30°C overnight.
-Incubate pRED at 30°C because of temperature sensitive promoter
-<br>
-pNPTS138-R6KT is for knock-ins via RecA Recombineering
 </td>
      </tr>
   <tr>
        <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
-       <td>No colonies. we suspected electroporation to be a problem and tried chemical transformation of NEB Turbo next.</td>
+       <td>No colonies. We suspected electroporation to be a problem and switched to chemical transformation in NEB Turbo next.</td>
      </tr>
 </table>
@@ Line 50: / Line 48: @@
   <tr>
        <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
-       <td>No colonies. We suspected the E. Coli NEB Turbo to be a problem and switched to E. Coli Dh5a as a cloning organism.</td>
+       <td>No colonies. We suspected the E. Coli NEB Turbo to be a problem and switched to E. Coli Dh5α.</td>
      </tr>
 </table>
-<h4>Redo: Transforming E.Coli Dh5a to amplify plasmids for pRED/ET Engineering</h4>
+<h4>Redo: Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering</h4>
 <em>2018/05/24</em>
 <table class="table table-borderless">
@@ Line 65: / Line 63: @@
        <td>Protocol:</td>
        <td>
-<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electrocompetent transformation</a>
+<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electro-transformation</a>
 </td>
      </tr>
@@ Line 74: / Line 72: @@
   <tr>
        <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
-       <td>No colonies</td>
+       <td>We got cells containing the plasmid from PD Dr. Jürgen Lassak of the LMU.</td>
      </tr>
 </table>
-<h4>Transforming E.Coli Dh5a to amplify plasmids for pRED/ET Engineering</h4>
+<h4>Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering</h4>
 <em>2018/05/25</em>
 <table class="table table-borderless">
@@ Line 113: / Line 111: @@
        <td>Protocol:</td>
        <td>
-<a href="https://international.neb.com/protocols/2015/11/20/monarch-plasmid-dna-miniprep-kit-protocol-t1010" target="_blank">Mini Prep</a>
+<a href="https://international.neb.com/protocols/2015/11/20/monarch-plasmid-dna-miniprep-kit-protocol-t1010" target="_blank">Miniprep</a>
 </td>
      </tr>
      <tr>
        <td>Notes:</td>
-       <td>because pRED/ET could not be transformed, we got readily transformed cells from PD Dr. Jürgen Lassak from the LMU</td>
+       <td>The cells from PD Dr. Jürgen Lassak were used for DNA preparation.</td>
      </tr>
   <tr>
        <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
-       <td>pRED/ET: 		37,5 ng/µl
+       <td>Concentration of the plasmids obtains from MIniprep: pRED/ET: 		37,5 ng/µl
-pNPTS138-R6KT:	60ng/ul
+pNPTS138-R6KT:	60ng/µl
 </td>
      </tr>