Team:DTU-Denmark/Notebook/28 August 2013
From 2013.igem.org
28 August 2013
Contents |
Lab 208
Main purpose
Who was in the lab
Henrike, Kristian, Julia
Procedure
A gradient PCR was made for pSB1C3 and pZA21::ara for Nir.
The master mix contains:
- CG buffer,
- X7 polymerase,
- 2% of DMSO (1uL of 100%DMSO) per reaction
- 1mM MgCl2 (1uL of 50mM MgCl2) per reaction
1 | 2 | |
---|---|---|
Template | pSB1C3 | pZA21::ara |
Primers | 57a + 57b | 58a + 58b |
Temperature range | 58C-78C | 50C-65C |
Elongation time per cycle | 1:30 min | 2 min |
Each reaction was run in one of the 12 different annealing temperatures within the range indicated in the table. We followed the standard PCR program.
Results
Conclusion
lab 115
Main purpose
Run Experiment 4 in two different samples anaerobically in order to characterize the behavior of AMO transformants.
Who was in the lab
Ariadni, Helen, Kashia
Procedure
Adjusting the temperature at 36 degrees and calibrating the probes as described in Calibration protocol.
Following the protocol Experiment 4
Changing the steps :
step 2 . 4 ml of the overnight culture growing in DM minimal medium with NH4Cl
and step 3 from the experimental procedure where the OD was measured OD=0.0761 and 0.0720 for the AMO mutant. For the E.coli culture the OD was 0.0822 and 0.0947.The abiotic controls have OD= 0.002 and 0.0006.
Results
Colorimetric results
Ranges
- Measuring range 5-150 mg/L NH4-N
- Measuring range 0.02-1 mg/L NO2-N
Standard solutions
- Ammonium - 66.6 mg/L (expected 39 mg/L)
- Nitrite - 0.72 mg/L (expected 0.5 mg/L)
time (min) | ammonium AMO1(mg/L) | ammonium AMO2(mg/L) | Ammonium E.coli(mg/L) | nitrite E.coli | ammonium abiotic (mg/L) | nitrite abiotic(mg/L) |
---|---|---|---|---|---|---|
0 | <5 | <5 | <5 | <0.02 | 5 | <0.02 |
13 | <5 | <5 | 5 | <0.02 | - | - |
19 | <5 | <5 | 5 | <0.02 | - | - |
spike | 146 | 140 | <5 | <0.02 | - | - |
35 | 158 | 244 | 258 | <0.02 | - | - |
41 | 169.5 | 171 | 178 | <0.02 | - | - |
52 | 226 | 280 | 250 | <0.02 | - | - |
64 | 232 | 254 | 124 | <0.02 | - | - |
90 | 238 | 272 | 246 | <0.02 | - | - |
10 hours | 172 | 218 | 16 | <0.02 | 154 | <0.02 |
OD at the end of the experiment
OD=0.2083 and 0.2232 for the AMO mutants. For the E.coli culture the OD was 2.39 and 0.1173.The abiotic controls have OD=0.00144 and 0.0004.
Conclusion
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