Team:Caltech/Notebook

From 2014.igem.org

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<ul><li>Linear constructs and plasmid TXTL reactions of A90 promoter (pAA001) were set up and run</li>
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     <li>Gibson assembly of plasmid pAA002 containing B83 promoter</li>
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     <li>PCR of Gibson assemblies of pAA002 to create linear constructs to test in TXTL</li>
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    <li>TXTL reactions of B83 promoter (linear frag. from pAA002) were set up and run</li>
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    <li>Transformation of pAA002 Gibson assemblies into JM109</li>
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<ul><li>Analysis of yesterday's TXTL reactions (run overnight)</li>
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     <li>Colony PCR and miniprep of bacterial colonies transformed with pAA002. Minipreps shipped for sequencing.</li>
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<td>Colonies were found and picked after transformation and incubation.</td>
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<td>TXTL reaction data inconclusive. Most fluorescence signals are below negative control, and linear data frequently contradicts plasmid data with A90 promoter. No signal observed for B83 promoter.</td>
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Revision as of 09:13, 27 June 2014



WELCOME TO iGEM 2014!

Your team has been approved and you are ready to start the iGEM season!
On this page you can document your project, introduce your team members, document your progress
and share your iGEM experience with the rest of the world!


Click here to edit this page!

Home Team Official Team Profile Project Parts Modeling Notebook Safety Attributions

Notebook

Apparently there's a calendar feature to take care of this too???

Week One

Tuesday, 6/17/14
  • Get acquainted with summer logistics and lab basics
  • Clean up lab space, stock lab supplies
 We got absolutely no results today!!!

Wednesday, 6/18/14
  • Continue to discuss project and clean up lab space
  • Designed PCR primers for combinatorial promoters subproject
 We got absolutely no results today!!!

Thursday, 6/19/14
  • Formulated plasmid design (pAA001) for the response regulation subproject
  • PCR run on plasmid backbone and sfGFP to create constructs with the proper overhangs for Gibson assembly
  • Gel electrophoresis for confirmation of PCR products' length
 With the exception of the lacI gene, PCR products were confirmed via gel electrophoresis

Friday, 6/20/14
  • Purification of PCR products created yesterday
  • Gibson assembly of purified products
  • Transformation of Gibson-assembled plasmids into JM109 E. coli
  • Meeting with Prof. Murray to discuss give an update on the project
 Colonies were found and picked after transformation and incubation.

Week Two

Monday, 6/23/14
  • Discussion of fallout from Friday's meeting, paper reading
  • Colony PCR and Minipreps of liquid cultures of colonies picked over the weekend
  • TXTL workshop starts today

Tuesday, 6/24/14
  • Discussed alternatives to current ComQXPA system, including 2-cell system (not all in 1 cell)
  • Gibson assembly of another combinatorial promoter using pKS001 backbone, lacI geneblock, ____ promoter geneblock, and sfGFP geneblock.
  • Transformation of Gibson constructs into JM109 cells
  • Linear combinatorial constructs created via PCR of pAA001 (minipreps) to be tested in TXTL

Wednesday, 6/25/14
  • Linear constructs and plasmid TXTL reactions of A90 promoter (pAA001) were set up and run
  • Gibson assembly of plasmid pAA002 containing B83 promoter
  • PCR of Gibson assemblies of pAA002 to create linear constructs to test in TXTL
  • TXTL reactions of B83 promoter (linear frag. from pAA002) were set up and run
  • Transformation of pAA002 Gibson assemblies into JM109

Thursday, 6/26/14
  • Analysis of yesterday's TXTL reactions (run overnight)
  • Colony PCR and miniprep of bacterial colonies transformed with pAA002. Minipreps shipped for sequencing.
 TXTL reaction data inconclusive. Most fluorescence signals are below negative control, and linear data frequently contradicts plasmid data with A90 promoter. No signal observed for B83 promoter.


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