Week 3 laboratory reflection - Isolation and Quantification of Microorganisms

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Indiana University, Bloomington *

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Biology

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Dec 6, 2023

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Name: Laboratory Reflection: Isolation and Quantification of Microorganisms Instructions: Make sure to scroll through the entire document to answer all questions. It would also help with grading if you put your typed answers in a different (but readable) color font . Save and upload your completed assignment to the associated assignment page on Canvas. It is the student’s responsibility to confirm that the correct file was submitted to Canvas by the due date. 1. (1pt) What is aseptic technique and why is it important? Aseptic technique is what we use to ensure clean samples when growing cultures. It is important to ensure accurate results, where no other cultures grow. 2. (1pt) List two aseptic technique procedures you learned about this week and describe each of their importance. For example, handwashing is important to reduce the number of microbes on the hands. Flame sterilizing the loop is important to ensure no cross contamination, and sterilizing all working surfaces ensures the same thing. 3. (1pt) When plating a microorganism, with the goal of quantifying specific colonies, why is performing a dilution necessary? If there are too many cultures on a Petri dish, the sample will be too difficult to count to find an accurate result. 4. Imagine you have been given a liquid culture of yeast with a starting concentration of 7.81 x 10 8 cells/ml and are asked to carry out the sample dilution process shown in the figure below. Based on this dilution scheme, answer questions a-e. Pay special attention to the units.

a. (1pt) You measured 100 µl of original yeast culture and used it to make your 10 -1 dilution (or 1/10 th dilution). How many yeast cells were present in this 100µl of original yeast culture? Show your work (make sure you include all units in your calculations). There would be 7.81x10^7, as it is dividing by 10, so I subtract 1 from the exponent. b. (1pt) The 100µl of original culture was added to 900µl of water to make the 10 -1 dilution. What was the concentration (in units of cells/ml) of the 10 -1 dilution? Show your work. This would have 10x less per mL than the first, so there would be 7.81x10^7 cells/mL c. (1pt) What was the concentration (in units of cells/ml) of the 10 -5 dilution? Show your work. There would be 7.81x10^3 cells/mL, as when dividing in dilution you would subtract the exponents, 8-5=3 d. (1pt) How many colonies should have been present on Plate A in this example? ( HINT: Calculate the number of cells that were present in the 100µl of solution from the 10 -3 dilution that you plated on Plate A; assuming that all of these cells grow into colonies, this should be the same as the number of colonies you count on the plate ). Show your work. 7.81x10^4, the whole solution has 7.81x10^5, but we are pipetting 10% of that to the plate. e. (1pt) How many colonies should have been present on Plate C? 7.81x10^2 1. (2pt) Imagine you carried out the same dilution scheme shown in the figure from question 4, but now, you do not know the concentration of the original culture. If you counted 344 colonies on Plate B using the image above, what is the concentration of cells/ml in the original culture? How many colonies would you expect to count on Plate A? Show your work and remember to pay attention to units.

3.44x10^6 cells/mL in the original culture, plate A would have 3440 colonies. We have to multiply by 10 4 times, as well as the 2 from going from 344 to 3.44.

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