Difference between revisions of "Part:BBa K1065002"

Figure 1 Effect of EFE on cell growth. Cell density was measured at different time points to determine the effect of EFE expression. Neb10β cells were grown at 37 °C in LB until an OD of 0.6. The cultures were then split in four samples of equal volume, and two samples were induced with 5 mM Arabinose. Induced samples show a slowed growth rate. This was expected as 5mM arabinose is considered a strong induction that causes moderate stress on cells.

Figure 2 Ethylene detection using an Agilent 3000 Micro GC set up with a plot U column. Neb10β cells were grown until an OD of 0.5. The culture was then split in two samples of equal volume (3 ml) and placed into an hermetically closed vial with a septum and a rubber cap. One of the two samples was induced with 5 mM arabinose. The vials were kept at 37°C under shaking for 4 hours. The vials were then connected to the Micro GC and measurements were performed. Panel A: induced 1.5 ml sample (green curve) and 3 ml sample (red curve) showed a characteristic peak corresponding to Ethylene. On the other hand, the 3 ml non-induced sample (blue curve) didn't show the Ethylene peak. Ethylene concentration was estimated to be 61 ± 15 ppm for the 1.5 ml culture and 101 ± 15 ppm for the 3 ml culture. Panel B: picture of the vial connected to the micro GC.

Figure 3 Kinetic assay for Ethylene production. Neb10β cells were grown in agitation on a thermostatic block at 37 °C until an OD of 0.5 - 0.8 and then connected to the Micro GC. Every 45-60 min a measure was taken for a total experiment time of about 8 hours. Samples were induced at two different OD and this had a marked effect on the amount of Ethylene produced. However, it seems that the Ethylene concentration in the air volume reached saturation after only two hours. The red dashed line indicates the amount of Ethylene detected with a culture left in the shaking incubator for the entire duration of the experiment and subjected to only one measurement. As expected, an higher concentration of Ethylene was measured due to the minimal gas loss using this approach.

Figure 4 Acceleration of fruit ripening. Panel A: our system was applied to the acceleration of fruit ripening. We designed hermetically closed jars with a rubber hose connector. These jars contained our test-fruit and each one was connected to a flask. The flasks contained 300 ml of induced (or non-induced) culture at an OD of 0.8. The flasks contained cultures were maintained at 37°C using a laboratory heating plate with a magnetic stirrer. During four to six days, cultures were substituted on a daily basis with a new induced (or non-induced) culture. Furthermore, non-modified jars (i.e. with no connector) were used for the negative control fruit samples (no cells sample). Panel B: ripening of plums. Plums exposed to Ethylene show a more advanced stage of ripening after 4 days of treatment respect to the negative controls (no cells and non- induced BBa_K1065001).

Figure 5 Neb10β cells transformed with BBa_K1065311: pellets after induction time. Cultures at an OD of 0.7 were exposed to a blue LED (470 nm) for about 10 hours (2). Control sample (1) was covered with an aluminum foil and maintained in complete darkness. A substantial difference between pellets' coloration can be observed. AmilCP production probably indicates that also Ethylene would be produced.