Difference between revisions of "Team:LambertGA/Notebook"

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<center> <h1 id="MainTitle"><b> Notebook </b></h1> </center>
 
<center> <h1 id="MainTitle"><b> Notebook </b></h1> </center>
  
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Week 1 (Aug 8):
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<br>
 
<br>
 
Safety procedures and aseptic technique was reviewed by our iGEM supervisor: Janet Standeven.
 
Safety procedures and aseptic technique was reviewed by our iGEM supervisor: Janet Standeven.
 
<br><br>
 
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Week 2 (Aug 15):
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This week was subjected to ligating two of our three parts of the genetic construct together: p-lambda-r/Lac I/ tspurple/ deg. Tag (LAA or DAS tags) + ROO1/ ClpXP/ CI
 
This week was subjected to ligating two of our three parts of the genetic construct together: p-lambda-r/Lac I/ tspurple/ deg. Tag (LAA or DAS tags) + ROO1/ ClpXP/ CI
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After ligation was complete the the genetic constructs were then transformed into separate DH10 <i>E. coli</i> cells  
 
After ligation was complete the the genetic constructs were then transformed into separate DH10 <i>E. coli</i> cells  
 
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Week 3 (Aug 22):  
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The previous transformation and ligation was unsuccessful due to the lack of ribosomal binding sites in the genetic constructs  
 
The previous transformation and ligation was unsuccessful due to the lack of ribosomal binding sites in the genetic constructs  
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Transformation and ligation of the third part: genetic construct without a degradation tag present  
 
Transformation and ligation of the third part: genetic construct without a degradation tag present  
 
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Week 4 (Aug 29):
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The continuation of the re-ligations and re-transformations during week 3 were ultimately unsuccessful due to RFP( red fluorescent protein)  contamination
 
The continuation of the re-ligations and re-transformations during week 3 were ultimately unsuccessful due to RFP( red fluorescent protein)  contamination
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Miniprep and nanodrop of the genetic construct without a present degradation tag in the backbone: 1C3.
 
Miniprep and nanodrop of the genetic construct without a present degradation tag in the backbone: 1C3.
 
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Week 5 (Sep 5):  
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Further detection of previous errors  
 
Further detection of previous errors  
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The above ligations were also transformed  
 
The above ligations were also transformed  
 
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Week 6 (Sep 12)
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Week 7 (Sep 19)
 
Week 7 (Sep 19)
 
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Week 8 (Sep 26)
 
Week 8 (Sep 26)
 
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Week 9 (Oct 3)
 
Week 9 (Oct 3)
 
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Week 10 (Oct 10)
 
Week 10 (Oct 10)
 
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Ligation  
 
Ligation  
 
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Revision as of 02:04, 16 October 2016



Notebook


Safety procedures and aseptic technique was reviewed by our iGEM supervisor: Janet Standeven.


This week was subjected to ligating two of our three parts of the genetic construct together: p-lambda-r/Lac I/ tspurple/ deg. Tag (LAA or DAS tags) + ROO1/ ClpXP/ CI
Note: no construct was ligated without a present degradation tag
After ligation was complete the the genetic constructs were then transformed into separate DH10 E. coli cells


The previous transformation and ligation was unsuccessful due to the lack of ribosomal binding sites in the genetic constructs
We were required to re-digest, re-transform and re-ligate the p-lambda-r/ LacI/ TsPurple/ deg.tag(LAA or DAS) + ROO11/ ClpXP/ CI
Transformation and ligation of the third part: genetic construct without a degradation tag present


The continuation of the re-ligations and re-transformations during week 3 were ultimately unsuccessful due to RFP( red fluorescent protein) contamination
Miniprep and nanodrop of the genetic construct without a present degradation tag in the backbone: 1C3.


Further detection of previous errors
Liquid culture of genetic construct without present degradation tag
Inoculation of RFP and previous TS purple colonies
Due to several unsuccessful procedures, we decide to religate p-lamba-r/LacI + ts purple (deg tag) and R0011/ClpXP + CI
The above ligations were also transformed



Week 7 (Sep 19)

Week 8 (Sep 26)

Week 9 (Oct 3)

Week 10 (Oct 10)
Successful transformation of TsPurple and GFP constructs with variation of degradations tag (LAA, DAS and no degradation tag) and IPTG levels ( 1 uM, 10 uM, 100 Um, 1 mM)
Ligation