NOTE: In this experiment, you will measure the rate of cellular respiration for germinating seeds by measuring the rate of exchange for oxygen. As oxygen is consumed to provide energy, germinating seeds release carbon dioxide. This carbon dioxide is absorbed by potassium carbonate and thus the overall gaseous pressure of the respirometer will be reduced. HYPOTHESES: The experimental hypothesis is that germinating seeds will show a greater rate of respiration than control glass beads. Additionally, that at higher temperatures, the rate of cellular respiration in the seeds will increase. The null hypotheses are that both glass beads and germinating seeds will display a similar rate of cellular respiration and that temperature has no effect on respiration.
To set up the control, first remove the plunger from a prepared microrespirometer and label it with a “CR” or “CH” respectively, depending on whether it is the control room temperature or heated treatment.
Add the glass microbeads to each of the microrespirometers up to the 0.5 mL mark, and push the plunger in the syringe to the 1 mL mark.
Place 3 – 4 washers onto the flared end of each microrespirometer to weigh them down while in the water bath. NOTE: The microrespirometer will sit capillary tube end up.
Set the control devices to the side.
To set up the experimental seated respirometers, remove the plunger from a prepared microrespirometer and label it with “ER” or “EH” respectively, depending on whether it is the experimental room temperature or heated treatment.
Measure 0.5 mL of germinating seeds into the tuberculin syringe of the microrespirometer.
Then, push the syringe to the 1 mL mark.
Place 3 – 4 washers onto the flared end of each microrespirometer to weigh them down while in the water bath.
To use the microrespirometers, carefully press the “CR” and “ER” microrespirometers into the room temperature water bath. The entire syringe should be submerged, but the capillary tubes should be entirely exposed. Remove water from the bath until this position is achieved. Be sure the top of the capillary tube is open.
Perform the same procedure (step 9) for the “CH” and “EH” microrespirometers to set up the higher temperature experimental group.
Wait 5 minutes before moving on to allow the temperature in the microrespirometers to equalize with the water bath.
Add a single drop of manometer fluid to the top of each of the four capillary tubes which seals the microrespirometer chambers. NOTE: If the chambers are working properly, the manometer fluid will be sucked into the capillary chamber.
Use the plunger set in the bath to suck the manometer fluid into the capillary. Stop when the fluid is about halfway down the capillary tube. IMPORTANT: The microrespirometers should not be disturbed once they are added to the bath. Do not bump the table or insert or remove anything to or from the water bath once the chambers have been sealed and equalized.
Mark the bottom edge of the manometer fluid that is closer to the chamber. This represents time point zero.
Set a timer to sound in 5 minutes.
After 5 minutes, create a new mark where the manometer fluid is on each microrespirometer. Repeat every 5 minutes until 25 minutes has passed or until the manometer fluid has traveled the entire length of the capillary tube.
Then, remove the respirometers from the water baths.
During clean up, manometers should either be deconstructed to be reused or disposed of in the trash.
