Enumerating Bacterial Culture by Spread Plate Method

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Enumerating Bacterial Culture by Spread Plate Method Questions: 1) What do microorganisms need to grow? 2) What does counting colonies tell us about the bacterial populations? Why does counting colonies work? 3) What are the limitations of a direct count of colonies? Lab procedure: 1) Put on gloves. 2) Finger vortex the bacterial culture tube you are given by the instructor to ensure complete mixing. 3) Aseptically transfer 100ul of the culture to the middle of a pre-warmed agar plate with a re-pipettor and yellow tip (be careful not to stab the tip into the agar). 4) Dispose of the contaminated pipet tip in an orange biohazard bag by using the eject button on the pipettor. 5) Aseptically unwrap a disposable plate spreader and spread the 100ul inoculation evenly over the surface of the Petri dish as demonstrated by the instructor. 6) Aseptically place the lid back on the Petri dish and allow it to rest for 15 minutes undisturbed. 7) Place the contaminated plate spreader back in its plastic sleave and dispose in an
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Unformatted text preview: orange biohazard bag. 8) After a 15 minute hold time on the bench, invert the Petri dish and place in a 37°C incubator (plates may be stacked to a maximum of 3 high). 9) Remove gloves and wash hands before leaving the lab. 10) Incubate for 24+/-2 hours at 37°C. 11) Put on gloves 12) Remove Petri dish from incubator and place it agar side down on the back-lit plate counter. Remove the lid from the Petri dish and adjust the magnifying lens so that you can see the colonies clearly. 13) Count all of the colonies (multiple colonies that are touching each other are counted as one colony). 14) Record your count in your notebook and take to the next class meeting. 15) Calculate the original bacterial density using the following equation: CFU/mL= (plate count/0.100mL)/dilution factor Example: (150 colonies / 0.100mL) / 10-6 = 1.5 x10 9...
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