Imacarae Week 6

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Assignment Shared Entries Individual Entries
Week 1 Class Journal Week 1 ----
Week 2 Class Journal Week 2 Imacarae Week 2
Week 3 Class Journal Week 3 HSF1/YGL073W Week 3
Week 4 Class Journal Week 4 Imacarae Week 4
Week 5 Class Journal Week 5 CancerSEA Week 5
Week 6 Class Journal Week 6 Imacarae Week 6
Week 7 Class Journal Week 7 Imacarae Week 7
Week 8 Class Journal Week 8 Imacarae Week 8
Week 9 Class Journal Week 9 Imacarae Week 9
Week 10 Class Journal Week 10 Imacarae Week 10
Week 11 Sulfiknights Imacarae Week 11
Week 12/13 Sulfiknights Sulfiknights DA Week 12/13
---- Sulfiknights Sulfiknights DA Week 14

Purpose

  • To learn what a DNA microassay is and to understand its relevance in various scientific fields.
  • To form a basic overview of how this is used in research so that we may employ similar techniques in our own research design.

Discovery Questions

  1. (Question 5, p. 110) Choose two genes from Figure 4.6b (PDF of figures on Brightspace) and draw a graph to represent the change in transcription over time.
    Imacarae gene transcr graph.png
    • This is the transcription for genes DIT1 and SPS100.
    • A dip below the x-axis shows folding regression while a rise above the x-axis shows in folding induction.
  2. (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.)
    • Gene X
      - 1 hour: black
      - 3 hour: bright red
      - 5 hour: black
      - 9 hour: dim green
    • Gene Y
      - 1 hour: black
      - 3 hour: bright red
      - 5 hour: dim green
      - 9 hour: bright green
    • Gene Z
      - 1 hour: black
      - 3 hour: medium red
      - 5 hour: bright red
      - 9 hour: bright red
  3. (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?
    • Yes, genes X and Y were transcribed similarly. They are both repressed and induced similarly at specific times.
  4. (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?
    • Most spots would be yellow at the first time point because this is the point in which the genes are being transcribed as two transcriptomes. The color comes from both equal amount of red and green tags.
  5. (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).
    • TEF4 was repressed.
    • TEF4's change in expression was most likely repressed in the absence of glucose so that cellular energy can be focussed on other needed functions. Without glucose, cells have limited amounts of available energy, so it will conserve energy and use the energy it has in areas where it is most needed for survival. The transcription of TEF4 is not necessary for cell survival, so energy will be directed away from this in an environment of limited glucose.
  6. (Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
    • The TCA, or Citric Acid Cycle, genes would be induced if glucose supply is running out because this cycle makes helps make ATP in the absence of glucose and oxygen. The cell would need to transcribe genes to encode for molecules that assist in this process so the organism can function.
  7. (Question 12, p. 120) What mechanism could the genome use to ensure genes for enzymes in a common pathway are induced or repressed simultaneously?
    • The guilt by association method can be used to help ensure that transcribed genes are regulated the same way. This can be done through DNA microassay techniques.
  8. (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?
    • Since TUP1 is a gene that encodes for proteins that repress glucose-repressed genes, so as TUP1 is deprived of glucose, the number of glucose-repressed genes will increase. Therefore, the spot color would get increasingly more red as time went on.
  9. (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?
    • Since it is the cells that express the Yap1 gene that are being suppressed by glucosed depletion, the production of Yap1 protein will decrease as well. Therefore, the spots of Yap1p will get more green as time goes on in the experiment.
  10. (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?
    • You would verify that you deleted TUP1 and overexpressed YAP1 if the spots of TUP1 were green and the spots of YAP1 were bright red at the end of the experiment.

Acknowledgements

  • To my Homework Partner, Iliana Crespin. We met during class and briefly discussed the assignment.
  • Except with what is noted above, this individual journal entry was completed by me and not copied from another source.

Imacarae (talk) 23:36, 8 October 2019 (PDT)

References

  • Reading material and questions by Campbell, A.M. and Heyer, L.J. (2003), “Chapter 4: Basic Research with DNA Microarrays”, in Discovering Genomics, Proteomics, and Bioinformatics, Cold Spring Harbor Laboratory Press, pp. 107-124. Retrieved October 8, 2019 from Brightspace.
  • TEF4 Background by SGD. (2019), TEF4/YKL081W Overview, in Saccharymyces Genome Database. Retrieved October 8, 2019 from [https://www.yeastgenome.org/locus/S000001564].
  • Assignment by LMU BioDB 2019. (2019). Week 6. Retrieved October 8, 2019 from [https://xmlpipedb.cs.lmu.edu/biodb/fall2019/index.php/Week_6].
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