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All of the previous sections have prepared you to grow the bacteria you desire. Since this section is quite long, we have broken it up into two main sections, growing bacteria for storage and growing bacteria for experiments.
As with all aspects of microbiology the aseptic technique is vital to your success.
PART 1: GROWING BACTERIA FOR STORAGE- plate-to-plate transfer
This is not for the permanent storage of bacteria, which will be discussed in another section. This is about the accessibility of the bacterial culture. When using bacteria on a daily basis, it is important to have the bacteria easily accessible. Bacteria is most accessible on a plate; therefore you must learn how to streak plates. The ability to effectively and efficiently streak plates is imperative to your success in the microbiology lab.
The first step in growing bacteria is transferring the bacteria into/onto the media which you desire. Most of the time, the microorganisms you buy from a company will be in a small vial, frozen. In order to allow these bacteria room to multiply, they must be transferred onto a media plate. A lab instructor will do this. One small vial of bacteria can last you a lifetime if the proper precautions are taken.
Once the bacteria has been transferred to a plate and grown, you will be able to see many isolated colonies. This is the type of plate you will often encounter. An isolated colony contains millions and millions of bacteria, which can be transferred to other media to be grown in colonies. This is referred to in the 'Growing bacteria for experiments' section. For now, we will just focus on growing bacteria for storage.
Bacteria, like everything else, gets old. It goes through lag, log, and stationary stages before death. We planed to include a graph of bacterial growth here but we changed out mind. In order for you to retain what you've learned better, you should go get your microbiology textbook now. From the index, find "growth curve." Now read the section on bacterial growth curve and write down the definitions of each of the following terms.
In the death phase, the bacteria will not perform up to the normal standards. In order to prevent using 'dead' bacteria, it is important to transfer the bacteria to a new plate. On this new plate the bacteria will have an opportunity to grow and multiply, starting over from the lag phase. Another reason for transferring bacteria to a new plate is that plates don't last forever. It will dry out and bacteria will eventually die. For these two obvious reasons, it is important to transfer the bacteria to a new plate at least every two months.
Streak a plate from an isolated colony on a plate. Use above instructions. Incubate. Count the number of isolated colonies. A good plate will have 10 isolated colonies. Continue practicing until you are comfortable transferring bacteria to plates and you are getting at least 10 isolated colonies.
PART 2: GROWING BACTERIA FOR USE IN EXPERIMENTS - plate-to-tube and tube-to-flask transfers.
A. Plate to tube transfer
WHY: A mass of bacteria is needed for most of the microbiology experiments you will be performing. Also, we need bacteria in a specific stage of their growth to ensure the consistency of our experiments and results. Therefore an isolated colony will not work. As mentioned before, an isolated colony has thousands of single bacterium in it. All of these bacteria are capable of multiplying and all of the multiplied bacteria are capable of multiplying also. After all this multiplying you will have a mass of bacteria. In order for the bacteria to multiply, the bacteria need nutrients and space. You must start small though. If one isolated colony were placed into a flask containing 500ml of media, the bacteria would be overwhelmed and die. If you place an isolated colony in a tube containing on only 5ml of media, the bacteria will flourish. Once the bacteria has flourished in the small tube, the entire contents of the tube can be transferred to the 500ml flask. Use the following steps as you transfer bacteria from a plate to a tube.
You should also note that if BEFORE you transfer the bacteria to the tube. If the tube is cloudy, there is a problem. DO NOT use that tube! The media is NOT sterile, and therefore your bacteria will be competing with other bacteria and it will not be optimal growing conditions for the bacteria you desire.
After the tube has been incubated it can be transferred to a larger flask in order to grow more bacteria. The tube CANNOT be kept in the refrigerator until a later date. Most of the time the amount of bacteria harvested from a 10ml tube is not enough for an experiment, therefore you will need to perform a tube to flask transfer.
B. Tube to flask transfer
WHY: A tube to flask transfer is necessary to grown a larger amount of bacteria. It won't work if you inoculate a colony-ful of bacteria from a plate directly to a big flask! The bacteria on a plate are used to growing on solid medium. You need to transfer them to a small amount of liquid medium so they can get used to liquid nutrient. Then they will be ready to grow in a larger amount of liquid medium.