Making GNA Plates
- First, grab a 250 mL flask and a 100 mL graduated cylinder from the glassware shelves (Shelf E and Shelf F). Make sure they have foil covering them and that the autoclave tape that is there is striped black.
- Gather a bag of scoopulas, a mass scale, and 4 medium-sized weighboats and place next to the hot plates on the bench.
- Using your graduated cylinder, obtain 100mL of dH2O and place it in the flask. Alongside this, place a small stir bar within the flask, but place it so that way you do not just drop it in the flask. You can do this by tilting the flask sideways a bit and sliding in the stir bar.
- Turn on the hot plate by only turning the stir knob, as you do not want the hot plate to be heating the flask while you are doing this.
- Within one weighboat and using one scoopula, gather 5g of D-Glucose. Make sure that when weighing the 5g of D-Glucose that the scale is tared with the weighboat so that your measurement is accurate. Add this to your flask.
- With another weighboat and scoopula, obtain 3g of Difco Nutrient Broth and add that to the flask as well.
- In your third weighboat, place 1g of Yeast Extract and in your fourth weighboat obtain 2g of Sunrise Agar. Add both of these substances to your flask.
- Make sure all material you added to the flask is well stirred and properly mixed. Once you are sure of this, place the beaker and a tub onto the cart. Put a new layer of tin foil covering the flask and place some autoclave tape on it.
- Take the cart with your items to either the first or second floor and push it to the room containing the autoclave.
- Once inside, put on your hot gloves (that should be on the cart) and open the autoclave if there is not a protocol running.
- Fill the tub with one inch of water and place it inside the autoclave on the top rack. Following this, place your flask of GNA mix in the water.
- Close the autoclave and make sure it is completely shut.
- Turn the autoclave onto the liquid cycle (You can do this by pressing the number 4 on it twice) and set a timer for 55 minutes. You can head back down to the lab and start cleaning up your area while the timer is running.
- Once the timer is done, go up to where the autoclave is and slightly open it. This will cause it to start to unload some pressure. The time for unloading should take 10 minutes, so it is best that you stay up there.
- Take out the GNA flask and put it onto your cart. Empty out the water into the sink located next to the autoclave. Be very careful while doing this, as the water is extremely hot and so is the metal on the autoclave. Make sure you are wearing your hot gloves the entire time you are doing this
- Take your GNA plate down to the lab and take four large plates and lay them out onto the bench. Grab a pipette aid and some large pipette tips.
- Using your pipette aid and the large pipette tips, pipette 25 mL of GNA into each large plate. Make sure to not move the plates to ensure that the solution solidifies properly.