The working method for doing these lab experiments were discussed by the modeller responsible for the Metabolic Model and the experimentalists.
Week 2
Experiment design and protocols were improved.
Week 3
Experiment design and protocols were improved.
Week 4
No lab work possible due to a medical issue.
September
Week 5
Experiment 1 from the E. coli survival protocol was done, but failed due to the 96 wells plate having been on a shaker in between steps, causing contamination of wells from other rows. The shaker setup can be seen in figure 1 and the result in figure 2.
Figure 1. Photo of plate 2 of the first experiment on the shaker. We believe this is the reason the contamination of Figure 2 occurred. Figure 2. Photo of plate 2 of the first experiment on a lightsource. As all rows are dark, including the ones that were supposed to be kept sterile, we can clearly see the experiment has failed.
Week 6
Experiment 1 was repeated, this time plate 2 was kept stationary in between inoculation steps. The new results can be seen in figure 3.
Figure 3. Photo of plate 2 of the repeat of the first experiment. The top row shows growth after having spent 30 minutes in the sugar water and then overnight growth. The time step increases for row 3 to 50 minutes, row 5 to 90 minutes, and row 7 to 120 minutes. Row 8 and all intermediate rows are negative controls. The first three columns are replicates of 0g/L sucrose, then 3 columns of 100g/L sucrose, then 3 columns of 200g/L sucrose, and finally 3 columns of 400g/L sucrose.
After a successful execution of experiment 1. The next phase was designed and executed, in the protocol this refers to the setup of Table 2. The results are seen in figure 4.
Figure 4. Photo of plate 2 of the second experiment. The top row shows growth after having spent 1 hour in the sugar water and then overnight growth. The time step increases for row 3 to 2 hours, etc (as shown in the right side of the photo). Row 8 and all intermediate rows are negative controls.The first 4 columns are replicates of 625g/L sucrose concentration, whilst the last 4 columns are replicates of the 312.5g/L sucrose concentration conditions.
After showing that the E. coli can survive for 6 hours in extremely high sucrose concentrations of 625g/L the experiment was continued in a newly prepared plate of LB medium overnight. The result of which can be seen in figure 5.
Figure 4. Photo of plate 2 of the third experiment. The top row shows growth after having spent at least 12 hours in the sugar water and then overnight growth. The first 4 columns are replicates of 625g/L sucrose concentration, whilst the last 4 columns are replicates of the 312.5g/L sucrose concentration conditions.
