Eyoung20 journal week 6
Contents
- 1 Purpose
- 2 Questions
- 2.1 (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. You can either create your plot in Excel and put the image up on your wiki page or you can do it by hand and upload a picture or scan.
- 2.2 (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.)
- 2.3 (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?
- 2.4 (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?
- 2.5 (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).
- 2.6 (Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
- 2.7 (Question 12, p. 120) What mechanism could the genome use to ensure genes for enzymes in a common pathway are induced or repressed simultaneously?
- 2.8 (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?
- 2.9 (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?
- 2.10 (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?
- 3 Acknowledgements
- 4 References
Purpose
This assignment is designed to give students an informative and active introduction to DNA microarrays, allowing students to gain a familiarity that will be useful in future projects.
Questions
(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. You can either create your plot in Excel and put the image up on your wiki page or you can do it by hand and upload a picture or scan.
This graph show the transcription of the genes DMC1 and SPS1. The data was gathered from figure 4.6 and interrupted using the fold repressed and induced values also found in figure 4.6.
(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 | 1 hour | 3 hour | 5 hour | 9 hour |
|---|---|---|---|---|
| gene x | 1.0 | 2.2 | 1.0 | 0.15 |
| gene Y | 1.0 | 4.5 | 0.95 | 0.05 |
| gene z | 1.0 | 1.5 | 2.0 | 2.0 |
| gene | 1 hour | 3 hour | 5 hour | 9 hour |
|---|---|---|---|---|
| gene x | Black | dim red | black | dim green |
| gene Y | Black | medium red | black | dim green |
| gene z | Black | dim red | dim red | dim red |
(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, gene x and gene y appear to have been transcribed similarly.
(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 a yellow spot is the result of no change occurring in the regulation of the gene. The yellow color is a combination of the green and red colors in equal parts. This yellow color is a result of no change in regulation due to a cell not being stressed, which is why the first point is yellow because the cell has not experienced the stressor yet.
(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 TEF4 was repressed as seen by the green color it has in Figure 4.12 at the time point labeled as OD 3.7. The cell mostlikly repressed the gene due to the reduction in glucose and the cell conserving energy reduced it's transcription levels.
(Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
The TCA cycle genes would be induced because the TCA cycle is a cycle that can be temporarily run without the presence of glucose, the use of this cycle will allow the cell to produce some energy while it starves. In essence it helps to keep the cells alive a little bit longer.
(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 genome uses shared promoters to control the the induction or repression of genes in common pathways.This means that by using one promoter the cell can control multiple genes and the production of enzymes.
(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?
You would expect the spots representing the glucose repressed genes to be very red, because TUP1 which is responsible for the repression these glucose repressed genes has been removed. Do to the removal there is nothing to stop or slow the expression of these genes meaning they will continue to increase in expression.
(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?
I would expect the spots of Yaplp targeted genes to be very red at later time points in the experiment. Yaplp is a transcription factor that aids in the resistance of environmental stress, so the longer experiment the continues, the more stress the cell is under and the more these genes should be expressed to attempt to compensate.
(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?
To confirm that you had truly deleted TUP1 you could run a microarray on cells that were provided with a surplus of glucose. The excess glucose would repress the glucose-repressed genes using TUP1 resulting in a green spot, however if TUP1 is removed then the spot will be yellow because there would be no change in the expression in the gene. The over expression of YAP1 would result in the the spots for the genes YAP1 promotes bing expressed and showing up as a red dot even if there is not stress applied to the samples that would promote the YAP1 stressor response.
Acknowledgements
I would like to acknowledge my homework partner user:Knguye66 for her help in understanding this assignment, and user:Kdahlquist for her instruction on the topic that provided valuable information and understanding.
"Except for what is noted above, this individual journal entry was completed by me and not copied from another source."
Eyoung20 (talk) 18:51, 9 October 2019 (PDT)
References
- Lodish, H., Berk, A., Zipursky, S. L., Matsudaira, P., Baltimore, D., & Darnell, J. (2000). Molecular cell biology 4th edition. National Center for Biotechnology Information, Bookshelf.
- Brown, P.O. & Botstein, D. (1999) Exploring the new world of the genome with DNA microarrays Nature Genetics 21: 33-37.
- 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. (Available on Brightspace)
- DeRisi, J.L., Iyer, V.R., and Brown, P.O. (1997) Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale. Science 278: 680-686.
- LMU BioDB 2019. (2019). Week 6. Retrieved October 9, 2019, from https://xmlpipedb.cs.lmu.edu/biodb/fall2019/index.php/Week_6
- Chapter 4 of Campbell & Heyer (2003)
