Testing Plant Substances as Possible Medicines
Purpose: The purpose of this lab is to find native plants in our areas of the field study, and see if they can help bacteria grow.
Materials: Balance, weigh boat, lab scoops LB broth base Media bottles, 250 mL Sterilize/autoclave Water bath, 37 degrees Celsius, shaking Sterile LB agar Laminar flow hood and disinfectant Glasses, safety, plastic Bunsen burner and gas lighter Inoculating loop, Ni/Cr wire Petri dishes, 60 x 15mm, 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 and filter, .2 microliters 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. Get mortar and pestle and grind 2 grams of plant with 10 mL of di H20. Then let it sit for 3 minutes. After this let this paste filter through a piece of paper that is lined with a funnel. Let this filter sterilize (.2 micrometers using syringe) and put it in a 1 mL microtube. 2. Repeat step 1, but instead of distilled water use 10 mL of Methanol. It is very toxic so use gloves and goggles.Put this extract in a heat block and leave the lid open to evaporate and leave it overnight. The next day add 1 mL of sterile water and vortex. 3. Sterilize forceps by putting it in flames, then put 2 discs in each of the extract and leave it overnight for the H20 extract or 15 minutes for the Methanol extract. 4. Once again get sterilized forceps and put 2 discs in each microbe with H20 (negative control). 5. Use flame to sterilize forceps once last time and put 1 disc in microtube with ampicillin (positive control). 6. Sterilize 1 mL pipet and transfer 1 mL of E. coli culture to a petri dish. Before you do that part divide the petri dish into 4 quadrants. Then spread the E. coli into all four quadrants with a sterilized glass spreader. Then cover the petri dish and let it soak for 15 minutes. 7. Sterilize forceps and place discs on the plates in each of the four quadrants s shown on the diagram below. 8. Incubate this plate at 37 degrees Celsius for 24 hours. 9. Measure qualitative and quantitative |