Background:
We went to Bio Marin and learned about different medicines and drugs they were creating. This lab is meant to expand on what we learned there and what we learned in class about medicine from plants.
Purpose/Objective:
What plant materials, found locally, contain active ingredients that will inhibit the growth of bacteria?
Materials:
Balance, weigh boat, lab scoops, LB broth base, media bottles, sterilizer, water bath, sterile LB agar, laminar flow hood and disinfectant, glasses, bunsen burner and gas lighter, inoculating loop, ni/cr wire, petri dishes, 60x15mm, sterile E. coli JM109 (stock plate) plant specimen, mortar and pestle, pipet, 10 mL and pump, plastic funnels, short-stemmed, filter paper disks, 5mm diameter beakers, 100mL syringe, 10 mL and filter, 0.2um reaction tubes and rack, 1.7 mL methanol, absolute pipet, 1 mL and pump dry block heater/heat block forceps, fine-tipped, ampicillin, glass spreader, incubator oven 37 degrees celsius
Procedure:
1.Measure 2 grams of plant material (lemon tree leaves)
2.Using a mortar and pestle, grind up 2 grams of plant tissue with 10 mL of deionized water
3.Let it sit for 3 minutes
4.Filter the sample through an 11 cm filter paper funnel
5.Filter sterilize the extract using a syringe filter
6.Collect 1 mL of extract into a 1.7 mL microtube. Label the tube
7.Repeat steps 1-6 using methanol instead of water
8.Attach pre-fitter to syringe and “rinse” with H2O9. Carefully open sterilization filter, keeping the filter in its plastic covering
10.Load approximately 1.5 mL of H2O-based filtrate using a pipet
11.Depress the plunger, collecting the sterile-filtered filtrate into microfuge tube
12.Quickly close microfuge cap until a “snap” sound is heard
13.Use Flame -Sterilized Forceps to transfer filter squares (2 per group) into H2O control and ampicalli and control microtubes
14.Label plate into four grids
15.Ms. Flasher will dispense 1 mL E-coli in each plate
16.Flame-Sterilize Spreading Loop and spread bacteria around
17. Close lid on both methanol and the water plates
Results:
We went to Bio Marin and learned about different medicines and drugs they were creating. This lab is meant to expand on what we learned there and what we learned in class about medicine from plants.
Purpose/Objective:
What plant materials, found locally, contain active ingredients that will inhibit the growth of bacteria?
Materials:
Balance, weigh boat, lab scoops, LB broth base, media bottles, sterilizer, water bath, sterile LB agar, laminar flow hood and disinfectant, glasses, bunsen burner and gas lighter, inoculating loop, ni/cr wire, petri dishes, 60x15mm, sterile E. coli JM109 (stock plate) plant specimen, mortar and pestle, pipet, 10 mL and pump, plastic funnels, short-stemmed, filter paper disks, 5mm diameter beakers, 100mL syringe, 10 mL and filter, 0.2um reaction tubes and rack, 1.7 mL methanol, absolute pipet, 1 mL and pump dry block heater/heat block forceps, fine-tipped, ampicillin, glass spreader, incubator oven 37 degrees celsius
Procedure:
1.Measure 2 grams of plant material (lemon tree leaves)
2.Using a mortar and pestle, grind up 2 grams of plant tissue with 10 mL of deionized water
3.Let it sit for 3 minutes
4.Filter the sample through an 11 cm filter paper funnel
5.Filter sterilize the extract using a syringe filter
6.Collect 1 mL of extract into a 1.7 mL microtube. Label the tube
7.Repeat steps 1-6 using methanol instead of water
8.Attach pre-fitter to syringe and “rinse” with H2O9. Carefully open sterilization filter, keeping the filter in its plastic covering
10.Load approximately 1.5 mL of H2O-based filtrate using a pipet
11.Depress the plunger, collecting the sterile-filtered filtrate into microfuge tube
12.Quickly close microfuge cap until a “snap” sound is heard
13.Use Flame -Sterilized Forceps to transfer filter squares (2 per group) into H2O control and ampicalli and control microtubes
14.Label plate into four grids
15.Ms. Flasher will dispense 1 mL E-coli in each plate
16.Flame-Sterilize Spreading Loop and spread bacteria around
17. Close lid on both methanol and the water plates
Results:
After 24 hours, we checked on our plates and saw no microbial action in either the methanol or water dishes except for the tiniest bit of clearance on the ampicillin portion of both the methanol and water dishes.
After 48 hours, we had a good amount of signs of contamination in the water dish and the ampicillin clearance had become more evident in the dishes on the ampicillin quarter.
After 72 hours, we saw the contamination had spread over the entirety of the water dish, and there are signs of contamination in the methanol. so basically we screwed up the whole lab, making us unable to fully analyze or even see our results.
After 48 hours, we had a good amount of signs of contamination in the water dish and the ampicillin clearance had become more evident in the dishes on the ampicillin quarter.
After 72 hours, we saw the contamination had spread over the entirety of the water dish, and there are signs of contamination in the methanol. so basically we screwed up the whole lab, making us unable to fully analyze or even see our results.
Analysis
Unfortunately, our dishes were contaminated. Once the dishes were contaminated the bacteria spread throughout our dishes and since there was so much bacteria in the dishes, we weren't able to correctly see our results. Luckily in the beginning, we were able to do the first step of the experiment, which was mash up the leaves and add different liquids such as de-ionized water, without problems. This lab really showed us the process of how a scientist could take any substance and test if it could be used as a potential medication to help treat a particular disease. As many scientists do, like I said before, we had some issues. We found that our dish had been contaminated so we were unable to find out what our plant had to offer. If we were to do this again, we would make sure our dish was air locked so now foreign bacteria could interfere with our results.