Dark vs. Light (Initial Conditions) Lab

Introduction

The purpose of this lab is to determine how initial light conditions of Halo cultures affect their growth in a lighted environment.

Hypothesis

The Halo in the dark environment will grow less than the Halo in the light environment because the Halo that was previously in the dark environment has to adjust in the light environment because it has to start producing bacteriorhodopsin, which it didn’t have in its dark environment in order to perform photosynthesis. On the other hand, the Halo in the light environment already was producing bacteriorhodopsin, so it already had the energy it needed to grow quickly.

Procedure:

1. Arrange six test tubes into two groups: three test tubes for light environment and three test tubes for the dark environment.
2. Using a spectrophotometer, calculate the OD of each test tube: Halo in the dark and Halo in the normal lighted environments.
3. Add 2.7 ml of Complete Media (CM) into each of the three test tubes designated for Halo in lighted environments.
4. Add 2.3 ml of Halo that were in a light environment into each of the three test tubes designated for Halo in lighted environments.
5. Add 3.6 ml of CM into each of the three test tubes designated for Halo in dark environments.
6. Add 1.4 ml of Halo that was in a dark environment into each of the three test tubes designated for Halo in dark environments.
7. For the control, add 50 ml of CM into one 125mL beaker.
8. Using a small 10 µl micropipette tip, pick up one Halo colony from the Falcon tube filled with Halo colonies and agarose. Drop the tip into the beaker with 50 ml of CM.
9. Make sure all lids of test tubes and beakers are fastened tightly.
10. Incubate at about 220 rpm and 37°C for 48 hours.

Results

Conclusion

After 72 hours, the light Halo, the Halo that initially came from a lighted environment, grew the most, having a final OD average of 1.177, while the dark Halo only grew to have an average OD of 0.263. It is possible that the dark Halo grew less because it was not in an active state, compared to the originally lighted Halo. Thus it took more time for the dark Halo to reach its log phase of growth and reproduce rapidly. In fact, since the dark Halo actually died during the course of the experiment, it is possible that the cells stayed dormant throughout the experiment and never grew at all. However, it is unclear whether the reason for the dark Halo’s dormant state was because of its dark environment or because of other external conditions, such as the amount of time it was kept on the shelf before the experiment. Therefore this experiment was inconclusive and should be repeated under more controlled conditions.