Introduction
This is a screening test used to determine whether gasoline-degrading bacteria are present. Gasoline is a relatively simple hydrocarbon so some bacteria should be able to use (degrade) it for their carbon and energy. In nature, it is likely that more than one species of bacteria is involved in degrading oil. Only one carbon source (gasoline) will be used and the other macronutrients (nitrogen, phosphorous, and sulfur) provided by the minimal salts.
One organism modifies an oil molecule so that another can attack it until the molecule is finally degraded. Degradation of the gasoline should result in growth on the plate containing minimal salts (nitrogen, phosphorous, sulfur, and magnesium) and gasoline as a carbon source.
OBJECTIVES
- Define bioremediation.
- To isolate gasoline-degrading bacteria from different environments.
METHODOLOGY
1. Soak a piece of filter paper or paper towel in the gasoline (10mL) and place the paper in the lid of one of the minimal-salts plates (1-2 drops of N & P fertilizers)
2. Place another gasoline-soaked paper in the lid of a water agar plate.
3. Place a water soaked paper towel in the lid of another minimal-salts plate. What is the purpose of each plate?
4. Pipette 0.51 mL soil slurries (a pinch of soil mixed in few mL of water) onto each of the agar plate.
5. Spread the soil over the surface of the agar with a L-rod.
6. Seal around the open edge of the plates with parafilm or a large rubber band to prevent escape of the gasoline.
7. Incubate the plates upside-down until colonies appear. Count the number of colonies on each plate.
RESULTS AND DISCUSSION
Guide Questions
- How could the gasoline-degrading bacteria you isolated be used to clean up a gas station or truck stop?
- How could you modify this experiment to isolate bacteria capable of degrading other hydrocarbons?
