Testing Plants
Purpose
To find which native plants have the correct ingredients to inhibit growth of bacteria.
Maretials
Plant specimen
10 mL syringe test tubes Methanol 1 mL pipet Ampicillin Inoculating Loop 60x15 Petri dish |
E. Coli JM109 (stock plate)
10 mL pipet 100 mL beakers 5mm filter paper 250 mL media bottle LB Agar LB broth base Plastic funnels |
Procedure
Part 1:
1. Prepare LB broth for E. coli. Wait 24 hours then add colony of E. coli to the broth.
2. Get 6 petri dishes and label them with your name and the date. Also, put a + sign on each dish then split the dish into 4 quarters.
3. Liquefy LB agar by heating it the microwave. Pour aprox. 20mL of agar into petri dish, then let agar solidify for 24 hours.
Part 2:
1. Grind up plant in deionized water. Then filter through filter paper then through syringe filter. Collect 1mL of this mixture and put it in a labeled test tube.
2. Repeat step 1 but use methanol instead of deionized water. Then place tube in a 65 celsius heat block with cap open for 24 hours. After, put 10mL of deionized water into the tube.
3. Repeat steps 2 and 3 until you have 6 samples. ( 3 of each)
4. Drop 3 filter disks into each tube.
5. Prepare 6 negative control disks of methanol and distilled water. (3 each)
6. Prepare 6 positive control disks of ampicillin.
7. Close tubes and store at 4 degrees Celsius.
Part 3:
1. Put 1mL of E.coli into your petri dish and spread around using a spreading loop. Wait for 15 minutes.
2. Place one disk in each quadrant using forceps. Be sure you have a positive and negative control disk for each sample.
3. Incubate the petri dishes at 37 degrees Celsius for 24 hours.
4. After 24 hours examine the petri dish fro zones of inhibition. Take pictures and record what you see.
1. Prepare LB broth for E. coli. Wait 24 hours then add colony of E. coli to the broth.
2. Get 6 petri dishes and label them with your name and the date. Also, put a + sign on each dish then split the dish into 4 quarters.
3. Liquefy LB agar by heating it the microwave. Pour aprox. 20mL of agar into petri dish, then let agar solidify for 24 hours.
Part 2:
1. Grind up plant in deionized water. Then filter through filter paper then through syringe filter. Collect 1mL of this mixture and put it in a labeled test tube.
2. Repeat step 1 but use methanol instead of deionized water. Then place tube in a 65 celsius heat block with cap open for 24 hours. After, put 10mL of deionized water into the tube.
3. Repeat steps 2 and 3 until you have 6 samples. ( 3 of each)
4. Drop 3 filter disks into each tube.
5. Prepare 6 negative control disks of methanol and distilled water. (3 each)
6. Prepare 6 positive control disks of ampicillin.
7. Close tubes and store at 4 degrees Celsius.
Part 3:
1. Put 1mL of E.coli into your petri dish and spread around using a spreading loop. Wait for 15 minutes.
2. Place one disk in each quadrant using forceps. Be sure you have a positive and negative control disk for each sample.
3. Incubate the petri dishes at 37 degrees Celsius for 24 hours.
4. After 24 hours examine the petri dish fro zones of inhibition. Take pictures and record what you see.
Results
Data Analysis
1. Did any extract give you positive results? Yes, the extracts both had positive results. 2. Did your controls work as expected? I thought my controls were going to be negative, but they were positive. 3. Discuss errors that could give you false results. Cross contamination, errors in pipetting, and incorrect use of the filter paper could have given us false results. 4. Discuss further experimentation. To further experiment, I would want to conduct this experiment on the rest of the plant species in my field study area. 5. Discuss next steps. To experiment on the rest of the plants, I would first need to collect the samples from my area. I have blackberries and bay trees in my area. I would take a leaf from each of the species and then follow the procedure. After doing this, I could conclude whether or not these other species are able to inhibit bacterial growth. |
conclusion
1. What did you like/find interesting?
I liked that on some of my plants I was able to to see white fuzzy splotches. I wasn't surprised that my plants were able to inhibit bacteria, but I was a little surprised because a lot of my plants were eaten away by caterpillars.
2. How did you and your partner collaborate?
My lab partner was Jason Franks. We worked together because we used the same plant, blackberry bushes. We collaborated well together and got the lab done in a timely manner.
3. What would you do differently next time?
I accidentally made a mistake in placing the disks in my petri dish. I accidentally put the wrong sample in the wrong quadrant. We also made another mistake and put to many filter disks in the tube.
I liked that on some of my plants I was able to to see white fuzzy splotches. I wasn't surprised that my plants were able to inhibit bacteria, but I was a little surprised because a lot of my plants were eaten away by caterpillars.
2. How did you and your partner collaborate?
My lab partner was Jason Franks. We worked together because we used the same plant, blackberry bushes. We collaborated well together and got the lab done in a timely manner.
3. What would you do differently next time?
I accidentally made a mistake in placing the disks in my petri dish. I accidentally put the wrong sample in the wrong quadrant. We also made another mistake and put to many filter disks in the tube.