replica plating

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University of Louisville *

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331

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Biology

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Apr 3, 2024

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pdf

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Uploaded by MajorMoon8437

Materials  LB and LB amp plates  Rubber Bands  250mL Flasks (or a better cylindrical object that fits the petri dishes)  Sharpies for labeling Procedure LAB: You will be provided 2 LB + amp plates, 1 LB plate, and one culture plate along with the materials to assemble your apparatus by the lab instructor. A. Clean the bottom of the conical component of the apparatus with 70% ethanol and let air dry. B. While conical component is drying, mark the LB + amp plates as 1 and 2 and the LB plate as “Control”. Make sure to mark a “North Point” on all of your plates and your velvet sheet as well. C. Once dry, assemble your replication apparatus. D. Place the apparatus gently into the culture plate on top of the agar surface, being careful to line up the North Points. E. Proceed to make two replicates by gently stamping both LB + amp plates 1 and 2, followed by the control plate. Place all three of your replicate plates together in an incubator set to 37 ˚C for 24 h. Place the culture plate into a 4 ˚C refrigerator for later comparison. THE DAY AFTER: Plates will be kept at 4 ˚C refrigerator following their incubation until the following lab. Sample Questions: Below are some example problems for Replica Plating: 1. If the replicates on the LB + amp plates follow Darwinian Evolution, what pattern would you expect to see in relation to the culture plate? You would expect to see the “survival of the fittest” principle enacted, where cells who had antibiotic resistance where then seen again on the second stamp. 2. If the replicates on the LB + amp plates follow Lamarckian Evolution, what pattern would you expect to see in relation to the culture plate? Lamarkian evolution describes how learned behaviors are passed onto offspring, and so, in the LB + amp plates you may see additional colonies that were produced as a result of learned antibiotic resistance.

3. Did you have any colonies missing on the control plates when compared to the culture plate? Were they present on the last LB plate as well? If you were to be missing colonies, what could explain their absence? The control group had a number of colonies missing when compared to the culture plate, which may be explained by them falling off during the stamping process. Therefore, if these missing colonies did not appear in the experimental plate, it can be attributed to errors in stamping rather than treatment with antibiotics. 4. According to your findings, whose hypothesis, Darwin, or Lamarck, explains the patterns seen on your plates. Darwin’s hypothesis of evolution is supported by our findings, as the number of colonies did not increase between plates. 5. What are the implications of these findings in regard to the evolution of antibiotic resistance in bacteria? This may help researchers to better understand how bacteria evolve and change overtime, which therefore may result in antibiotic resistance. Therefore, when developing antibiotic treatments for bacteria-caused diseases, we may properly respond to changes in bacterial behavior.

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