Lab 6d
Purpose: We wanted to find if our plants contain active ingredients that will inhibit the growth of bacteria.
Materials:
Balance, weigh boat, lab scoops
LB broth base
Media bottles, 250 mL
Sterilizer/autoclave
Water bath, 37 degrees Celsius, shaking
Sterile LB agar
Laminar flow hood and disinfectant
Glasses, safety, plasti
Bunsen burner and gas lighter
Inoculating 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 5mm diameter
Beakers, 100 mL
Syringe, 10 mL and filter, 0.2 um
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:
Part II: Preparing Plant Extracts
1. With a mortar and pestle, grind 2 grams of plant combined with 10 mL of deionized water. Let it sit for 3 minutes then let the paste filter through paper in a funnel. Let this filter sterilize (.2 micrometers using syringe) and put it in a 1 mL microtube.
2. Repeat step 1, but use 10 mL of methanol instead of deionized water. Use gloves and goggles to prevent burns from the methanol. Put these plant extracrs in a heat block and let it evaporate with the lid open. After evaporation add 1 mL of deionized water and mix it.
3. Put forceps in alcohol and burn them to sterilize. Put disks in each of the extracts and leave these overnight.
4. Use sterile forceps and put 2 disks in each microbe with water to make a positive control.
5. Use sterile forceps and put 1 disk in microtube with ampicillin to make a positive control.
6. Divide the petri dish into 4 quadrants labeling 2 water and 2 methanol with the positive and negative methanol labeling. Use sterile 1 mL pipet and transfer 1 mL of E. coli into a petri dish. Use glass spreader and spread the bacteria into all the quadrants and let it soak for about 15 minutes.
7. Use sterile forceps and place disks on the plates in each of the four quadrants.
8. Heat this plate at 37 degrees Celsius for one day.
Materials:
Balance, weigh boat, lab scoops
LB broth base
Media bottles, 250 mL
Sterilizer/autoclave
Water bath, 37 degrees Celsius, shaking
Sterile LB agar
Laminar flow hood and disinfectant
Glasses, safety, plasti
Bunsen burner and gas lighter
Inoculating 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 5mm diameter
Beakers, 100 mL
Syringe, 10 mL and filter, 0.2 um
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:
Part II: Preparing Plant Extracts
1. With a mortar and pestle, grind 2 grams of plant combined with 10 mL of deionized water. Let it sit for 3 minutes then let the paste filter through paper in a funnel. Let this filter sterilize (.2 micrometers using syringe) and put it in a 1 mL microtube.
2. Repeat step 1, but use 10 mL of methanol instead of deionized water. Use gloves and goggles to prevent burns from the methanol. Put these plant extracrs in a heat block and let it evaporate with the lid open. After evaporation add 1 mL of deionized water and mix it.
3. Put forceps in alcohol and burn them to sterilize. Put disks in each of the extracts and leave these overnight.
4. Use sterile forceps and put 2 disks in each microbe with water to make a positive control.
5. Use sterile forceps and put 1 disk in microtube with ampicillin to make a positive control.
6. Divide the petri dish into 4 quadrants labeling 2 water and 2 methanol with the positive and negative methanol labeling. Use sterile 1 mL pipet and transfer 1 mL of E. coli into a petri dish. Use glass spreader and spread the bacteria into all the quadrants and let it soak for about 15 minutes.
7. Use sterile forceps and place disks on the plates in each of the four quadrants.
8. Heat this plate at 37 degrees Celsius for one day.