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Team:KULeuven/14 August 2009
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Latest revision as of 08:47, 10 September 2009
Project progress
Progress of parts
[edit] Blue Light Receptor
- Transfer of the plates with cells from the ligation: 20 colonies have been transferred to a new plate to achieve more single colonies
- Plating of from -80°C
- Electroporation of the blue light promotor ligation (with ) in new competent cells
[edit] Vanillin Production
- Gel extraction of the 4 lanes from sam8 () and ech ()
- To test an additional blanc, we extracted a blanc piece of gel
- Nanodrop results were done with the conventional blanc (just elution buffer) and with the extracted blanco gel. However, since the results between both varied too much and the extracted blanco gel was not significantly better, we decided to use the conventional blanco in order to compare between former nanodrop measurements
Nanodrop:
| Part | concentration (ng/μl) | 260/280 λ | 260/230 λ |
|---|---|---|---|
| sam8 | 9,4 | 2,18 | |
| ech | 13,6 | 1,94 |
- Ligation of sam5 (from Wednesday) + sam8 (extracted today)
| vector | insert |
|---|---|
| sam5 | sam8 |
| 3637 bp | 1550 bp |
| 50 ng | 85,2 ng |
| 7 μl | 9 μl |
- Ligation of ech and fcs. For fcs we took the digest from last week (C1), because this week's restriction digest had a very low concentration (4,2) and a very high 260/280 (6,08)
| vector | insert |
|---|---|
| ech | fcs |
| 2929 bp | 1789 bp |
| 50 ng | 122 ng |
| 3 μl | 9 μl |
- Cells were stored at 16°C
- Plated the electroporated cells from the A and C ligation
- Made Agar and poured new plates with Ap
[edit] Vanillin Receptor
- G 2,3,1' on gel electrophoresis
- 2 and 3 seem oke, 1000bp
- 1' seems oke
- 800bp fragment + 200bp TOPO)
- Inoculate 2 and 3 and grow on ....
- TOPO from W and Z and again from A
- Note: if A fails again (with the iGEM protocol) then proceed with a new PCR purification.
- Mutagenesis 1 on position 202 of R was performed
