Purpose: To find what plant materials, found locally, contain active ingredients that will inhibit the growth of bacteria
Materials: Balance, weight boat, lab scoops, LB bath broth,media bottle 250mL, sterilizer/autoclave, water bath 37 degrees C, shaking sterile LB agar, Laminar flow hood and disinfectant, safety glasses, plastic bunsen burner, gas lighter, inoculatting loop, Ni/Cr wire. Petri dishes, 60x15 mm, sterile E. Coli JM109 (stock plate), Plant specimen, mortar and pestle, pipet, 10 mL and pump, plastic funnels, short stemmed, filter paper disks, 5 mm diameter, beakers, 100 mL, syringe, 10 mL filter, reaction tubes and rack, 1.7 mL, methonal, absolute pipet, 1 mL and pump,, dry block heater/heat block, forecps, fine-tipped, ampicillin, glass spreader, incubator oven 37 degrees C
Procedure:
Part 2
4. With a mortar and pestle, grind 2 g of plant tissue (leaves or bark) with 10 mL of deionized water until it turns into a paste. Let it sit for 3 minutes. Then, filter the paste through an 11 cm filter paper funnel. Filter sterilize the extract using a syringe filter. Collect 1 mL of extract into a 1.7 mL microtube. Label the sample.
5. repeat step 4, but using methanol instead of water. After, place the 1.7 mL tube with the 1 mL of methanol extract in 65 degrees C heat blocks for 24 hours
6. repeat steps 4 and 5 for all other samples
7. drop 3 filter paper disks into each tube of filtered extract using sterile forceps
8. prepare negative control disks, 3 each, of only methanol and sterile distilled water
9. prepare 6 positive control disks of ampicillin solution
10. allow the disks sufficient time to soak up enough extract to be saturated
11. close the tubes. Store all samples at 4 degrees C until ready to use.
Part 3
12. Using a sterile pipet transfer 1 mL of E. coli to middle of petri dish. Sterilize a spreader and evenly spread the bacteria around dish. Cover and let sit for 15 minutes
13. Using sterile forceps carefully place 3 disks into each quadrant. Keep all methanol and water samples on the same dish.
14. Place a positive control disk and a negative control disk either methanol or water into the appropriate quadrant
Data Analysis/Conclusion:
We got one positive result and 3 negative results (one of which was boarder line positive.) Our negative control worked as expected but our positive control was also negative. Something that could have given us false results was that we didn't label our samples with what they were, we just labeled them with our initials. We could have labeled the samples correctly and that probably would have fixed our problems that we ran into.
Materials: Balance, weight boat, lab scoops, LB bath broth,media bottle 250mL, sterilizer/autoclave, water bath 37 degrees C, shaking sterile LB agar, Laminar flow hood and disinfectant, safety glasses, plastic bunsen burner, gas lighter, inoculatting loop, Ni/Cr wire. Petri dishes, 60x15 mm, sterile E. Coli JM109 (stock plate), Plant specimen, mortar and pestle, pipet, 10 mL and pump, plastic funnels, short stemmed, filter paper disks, 5 mm diameter, beakers, 100 mL, syringe, 10 mL filter, reaction tubes and rack, 1.7 mL, methonal, absolute pipet, 1 mL and pump,, dry block heater/heat block, forecps, fine-tipped, ampicillin, glass spreader, incubator oven 37 degrees C
Procedure:
Part 2
4. With a mortar and pestle, grind 2 g of plant tissue (leaves or bark) with 10 mL of deionized water until it turns into a paste. Let it sit for 3 minutes. Then, filter the paste through an 11 cm filter paper funnel. Filter sterilize the extract using a syringe filter. Collect 1 mL of extract into a 1.7 mL microtube. Label the sample.
5. repeat step 4, but using methanol instead of water. After, place the 1.7 mL tube with the 1 mL of methanol extract in 65 degrees C heat blocks for 24 hours
6. repeat steps 4 and 5 for all other samples
7. drop 3 filter paper disks into each tube of filtered extract using sterile forceps
8. prepare negative control disks, 3 each, of only methanol and sterile distilled water
9. prepare 6 positive control disks of ampicillin solution
10. allow the disks sufficient time to soak up enough extract to be saturated
11. close the tubes. Store all samples at 4 degrees C until ready to use.
Part 3
12. Using a sterile pipet transfer 1 mL of E. coli to middle of petri dish. Sterilize a spreader and evenly spread the bacteria around dish. Cover and let sit for 15 minutes
13. Using sterile forceps carefully place 3 disks into each quadrant. Keep all methanol and water samples on the same dish.
14. Place a positive control disk and a negative control disk either methanol or water into the appropriate quadrant
Data Analysis/Conclusion:
We got one positive result and 3 negative results (one of which was boarder line positive.) Our negative control worked as expected but our positive control was also negative. Something that could have given us false results was that we didn't label our samples with what they were, we just labeled them with our initials. We could have labeled the samples correctly and that probably would have fixed our problems that we ran into.