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Team:DTU-Denmark/Notebook/10 July 2013
From 2013.igem.org
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* H<sub>2</sub>O: 31.5 ul (315 ul) | * H<sub>2</sub>O: 31.5 ul (315 ul) | ||
| - | Settings for PCR | + | Settings for PCR with three different elongation times |
{| class="wikitable sortable" style="text-align: right" | {| class="wikitable sortable" style="text-align: right" | ||
! Temperature (<sup>o</sup>C)!! Time (min)!! Rounds | ! Temperature (<sup>o</sup>C)!! Time (min)!! Rounds | ||
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| 66 || 0:45 || 35 | | 66 || 0:45 || 35 | ||
|- | |- | ||
| - | | 72 || 2:00 || 35 | + | | 72 || 2:00/3:00/4:00 || 35 |
|- | |- | ||
| 72 || 5:00 || 1 | | 72 || 5:00 || 1 | ||
Revision as of 16:23, 10 July 2013
Contents |
208
Main purpose
Run gel with 9 PCR samples of Nir operon from Pseudomonas aeruginosa
Transformation of Biobricks
PCR reaction for Nir operon
Who was in the lab
Ariadni,Henrike,Julia,Natalia
Procedure
Run gel
- 6 samples from PCR of 09.07.2013
- ladder broadband
- 3 samples from purification of 09.07.2013
Transformation of Biobricks
According to the iGEM protocol (transformation protocol) with slight changes.
- step 1: 100 μl cells
- step 2: 1.5 μl of the resuspended DNA
- step 6: 90 sec (instead of 60 s)at 42 oC
- step 9: Incubation without shaking
Extraction PCR
9 samples from culture pAO1 10 reactions
- dNTP: 1 ul (10 ul)
- HF buffer: 10 ul (100 ul)
- Phusion polymerase: 0.5 ul (5 ul)
- H2O: 31.5 ul (315 ul)
Settings for PCR with three different elongation times
| Temperature (oC) | Time (min) | Rounds |
|---|---|---|
| 98 | 2:00 | 1 |
| 98 | 0:20 | 35 |
| 66 | 0:45 | 35 |
| 72 | 2:00/3:00/4:00 | 35 |
| 72 | 5:00 | 1 |
| 4 | ∞ | - |
