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Team:TU-Eindhoven/Notebook/Week2
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Since all the BL21 pYES3-plates were empty, we have to <span class= "red"> redo the transformation</span>. 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. | Since all the BL21 pYES3-plates were empty, we have to <span class= "red"> redo the transformation</span>. 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 <span class= "red">fluorescence</span> of the GECO decreases again when the calcium concentration is increased. This is <span class= "red">contradictory</span> to what we expected. It seems that the GECOs we have in the freezer are already statured with calcium, so adding more CaCl<sub>2</sub> will | + | We determined the spectra with a Trisma buffer and concluded that the <span class= "red">fluorescence</span> of the GECO decreases again when the calcium concentration is increased. This is <span class= "red">contradictory</span> to what we expected. It seems that the GECOs we have in the freezer are already statured with calcium, so adding more CaCl<sub>2</sub> solution will only dilute the sample, after which <span class= "red">the fluorescence decreases</span>. 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. |
Revision as of 00:56, 27 September 2012
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 the fluorescence decreases. 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.
