Team:TU-Eindhoven/Notebook/Week2

• Introduction
  • Home page
  • Background
    information
  • About us
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• Project
  • Project description
  • Lab approach
  • Device information
  • Initial model
  • Yeast model
  • Lab results
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  • Our
    accomplishments
• Considerations
  • Public outreach
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• Future applications
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    developments
• Notebook
  • Before week 1
  • Week 1
  • Week 2
  • Week 3
  • Week 4
  • Week 5
  • Week 6
  • Week 7
  • Week 8
  • Week 9
  • Week 10
  • Week 11
  • Week 12
  • After week 12
  • Lab protocols
• Partners
  • ICMS TU/e
  • Team collaborations
  • Attributions and Contributions
  • Our sponsors

Things we did this week

The new mail address is ready to use (igem@tue.nl) for PR and sponsoring. After an interesting mail from the other Dutch iGEM teams, we went to a meeting in Amsterdam about the Discovery Festival. It's a festival where science meets art and is organized at Amsterdam, Rotterdam and Eindhoven. Since Friday, we have our own office in the renovated building 'Ceres' at the campus of the Eindhoven University of Technology.

Progression in the lab

Since all the BL21 pYES3-plates were empty, we have to redo the transformation. The one colony grown on the yeast pYES2-plates will be analyzed. The old E. coli containing R- and G-GECO proteins were cultured from the plates, so we can isolate the plasmids for long term storage. To find out whether the yeast contains the red GECO, a culture is put up. Furthermore, we decided to postpone the test of the device on the yeast cells.

We determined the spectra with a Trisma buffer and concluded that the fluorescence of the GECO decreases again when the calcium concentration is increased. This is contradictory to what we expected. It seems that the GECOs we have in the freezer are already statured with calcium, so adding more CaCl₂ solution will only dilute the sample, after which fluorescence decreases. A drawback of using CaCl₂ is that it affects the pH of the mixture. The solution is to purify the proteins again, this time by washing with EDTA to drain away the calcium and adding calcium through calcium acetate in a MOPS-buffer afterwards. Finally we revised the manual for determining GECO kinetics with a membrane based method for filtering out calcium from the isolated proteins.

Updating the software

In the device software, a square wave is added to support pulse trains. This can be applied to only one pixel now, but we will improve this in time.