Obtaining a Pure Culture from Isolated Colonies: Part 3

By TT -

 Introduction 

Previously, a hypothesis was tested that if a previously incubated 15mL tube with a tight lid was re-incubated, the bacterial cells would grow.  This test came out to be true, in this instance. Those bacterial cells were used to grow colonies on an agar plate. 

Material & Procedures

To culture the agar plate, the following items are needed: an agar plate, cultured liquid media (broth), a metal inoculation loop, a bunsen burner, a striker, and an incubator. 

First, install the bunsen burner to a gas source.  Once it is installed, turn on the gas, there should be a low hissing sound when the gas is released, and use the striker to ignite the flame. Now, take the inoculation loop and sterilize it using the flame allowing it to burn until it is glowing red, sterilize all parts that will enter into the container that contains the cultured broth.  Once it's ready, allow the loop to cool down, being mindful to keep it under the flame (next to the bunsen burner) to prevent contaminants in the air from attaching to the inoculation loop (about 20 seconds). After the cooling period, insert the loop into the container that holds the cultured broth, take the broth that is now on the loop, and spread it in the first section of the agar plate.  Next, sterilize the loop and repeat the cooling process.  From the first section drag cells into the next section of the agar plate to spread cells, repeat this process until cells are spread to grow colonies. Once the streaking process is complete, sterilize the inoculation loop and cut off the gas.  

To allow colonies the opportunity for growth, set in an incubator.  Deinococcus radiodurans should incubate at a temperature of 27°C. 

Results



After incubating the cultured agar dish, there were yellow colonies as opposed to orange colonies that are usually formed by D. radiodurans. This indicates that the culture had been contaminated. There are many potential causes of contamination, such as using improperly cleaned tools or contamination from the air due to not holding the loop under the flame when allowing it to cool. Contamination is suspected to be from an outside source, so, further examination of this culture will presume. 

Comments

  1. EricaMarch 30, 2022 at 5:01 PM

    Contamination happens, don't let it discourage you. Work on those aseptic techniques and you'll have it down in no time.

    ReplyDelete

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