Troque Week 7

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Introduction to DNA MicroArrays

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Q & A

Answer the following questions related to Chapter 4 of Campbell & Heyer (2003). Note that some of the questions below have been reworded from the Discovery Questions in the book:

  • (Question 5, p. 110) Choose two genes from Figure 4.6b (PDF of figures on MyLMUConnect) and draw a graph to represent the change in transcription over time. You can either create your plot in Excel and put the image up on your wiki page or you can do it in hard copy and turn it in in class.
TR transcription dmc1 sps1.png
  • (Question 6b, p. 110) Look at Figure 4.7, which depicts the loss of oxygen over time and the transcriptional response of three genes. These data are the ratios of transcription for genes X, Y, and Z during the depletion of oxygen. Using the color scale from Figure 4.6, determine the color for each ratio in Figure 4.7b. (Use the nomenclature "bright green", "medium green", "dim green", "black", "dim red", "medium red", or "bright red" for your answers.)
TR q2 table.png
  • (Question 7, p. 110) Were any of the genes in Figure 4.7b transcribed similarly? If so, which ones were transcribed similarly to which ones?
    • It would seem that genes X and Y are transcribed similarly. How I see it from the nomenclature I gave them in the table above is that, in the first hour, they both have no change in transcription. In hour 3, there is some noticeable change in their transcription; more specifically, there is inductive transcription (although it is much more noticeable in gene Y). For hours 5 and 9, there seems to be more repression in both as seen by the dropping numbers (although gene X's change is much slower as seen by the black color in hour 5 in contrast to gene Y's slightly lower ratio).
  • (Question 9, p. 118) Why would most spots be yellow at the first time point? I.e., what is the technical reason that spots show up as yellow - where does the yellow color come from? And, what would be the biological reason that the experiment resulted in most spots being yellow?
    • Yellow spots indicate that there is an equal amount of repression and induction. However, in this case, most spots would be yellow at the first time point since they would still need to adjust to the change in the environment. Repression/induction wouldn't happen in an instant so the genes are still at their baseline normal expression state.
  • (Question 10, p. 118) Go to the Saccharomyces Genome Database and search for the gene TEF4; you will see it is involved in translation. Look at the time point labeled OD 3.7 in Figure 4.12, and find the TEF4 spot. Over the course of this experiment, was TEF4 induced or repressed? Hypothesize why TEF4’s change in expression was part of the cell’s response to a reduction in available glucose (i.e., the only available food).
    • Over the course of the experiment, it would seem that TEF4 is slowly being repressed since it started out as yellow, but gradually began turning slightly greenish in color. A reduction in available food means that the cell has to adjust to the decreasing levels. I think that it would turn off the genes that are using glucose in order to compensate for the decreasing glucose levels, therefore, TEF4 expression is repressed.
  • (Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
    • TCA means tricarboxylic acid (cycle). I quick Google search reveals that this is "a series of enzyme-catalyzed chemical reactions that form a key part of aerobic respiration in cells" (Source: hyperphysics.com). This means that as the levels of glucose decreases, there has to be another way for the cells to produce food, this other way being the TCA cycle. According to the article, the TCA cycle induces some enzymes that are responsible for creating storage sugars form glucose-6-phosphate. The yeast cells were able to detect the decreasing glucose levels (thus, they know that they were about to starve), and were able to repress/induce certain genes in concert in order to continue their processes.
  • (Question 12, p. 120) What mechanism could the genome use to ensure genes for enzymes in a common pathway are induced or repressed simultaneously?
  • (Question 13, p. 121) Consider a microarray experiment where cells deleted for the repressor TUP1 were subjected to the same experiment of a timecourse of glucose depletion where cells at t0 (plenty of glucose available) are labeled green and cells at later timepoints (glucose depleted) are labeled red. What color would you expect the spots that represented glucose-repressed genes to be in the later time points of this experiment?
  • (Question 14, p. 121) Consider a microarray experiment where cells that overexpress the transcription factor Yap1p were subjected to the same experiment of a timecourse of glucose depletion where cells at t0 (plenty of glucose available) are labeled green and cells at later timepoints (glucose depleted) are labeled red. What color would you expect the spots that represented Yap1p target genes to be in the later time points of this experiment?
  • (Question 16, p. 121) Using the microarray data, how could you verify that you had truly deleted TUP1 or overexpressed YAP1 in the experiments described in questions 8 and 9?

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