Obtaining a Pure Culture from Isolated Colonies: Part 1

By TT -

 Introduction

Collecting a colony of bacteria from a petri dish is a process that can be used to produce a cultured broth.  Deinococcus radiodurans is an aerobic organism that was spread in a dish that contained a solid medium made with agar.  The bacteria, D. radiodurans, was not treated. So, the growth of this bacteria was expected to be normal with isolated colonies.  

Materials & Procedures

Materials that were used are as follows: A cultured dish with isolated colonies of D. radiodurans, 15 mL centrifuge tube, a TGY media (broth) in 500 mL Erlenmeyer flask, inoculation loop, a 10 mL pipette, a bunsen burner, nanometer, and an orbital floor shaker. 

There was 5mL of TGY (tryptone, glucose, and yeast extract) media placed in the 15 mL centrifuge tube using a 10 mL pipette. The bunsen burner was used to sterilize the lip of the Erlenmeyer flask before the TGY media was retrieved from the flask. After the media had been transferred to the tube, the inoculation loop was sterilized using the flame from the bunsen burner until it was glowingly hot.  After it was sterilized, it was allowed to cool for about 20-30 seconds, being held to avoid contact with any surfaces.  Then, the petri dish containing the isolated colonies of the D. radiodurans was taken and held upside down to avoid contaminates from the air and one colony was extracted from the dish and placed directly into the 15 mL tube that contained the 5 mL of media.  

After the colony had been added to the centrifuge tube the cap was placed on the tube and the tube was incubated at 27 degrees celsius in an orbital floor shaker for 48 hours. 

Results & Conclusions

After 48 hours the incubation process was ceased and there was little to no growth.  There was a small orange-colored pellet at the bottom of the tube.  The media was clear with a slight orangish tint.  A nanometer was used to measure the concentration or number of cells present using an OD600 reading.  The result was -0.06, indicating that there was no growth in the tube. This lack of growth could be from the lack of oxygen allowed to flow into the tube, hindering the growth process. This lack of growth could also be due to the lack of oxygen killing the bacteria resulting in no growth.  

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