Cdomin12 Week 11

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Assignment Page Individual Journal Entries Class Journal
Week 1 cdomin12 Week 1 Class Journal Week 1
Week 2 cdomin12 Week 2 Class Journal Week 2
Week 3 RAD53 / YPL153C Week 3 Class Journal Week 3
Week 4 cdomin12 Week 4 Class Journal Week 4
Week 5 IMG/VR Week 5 Class Journal Week 5
Week 6 cdomin12 Week 6 Class Journal Week 6
Week 7 cdomin12 Week 7 Class Journal Week 7
Week 8 cdomin12 Week 8 Class Journal Week 8
Week 9 cdomin12 Week 9 Class Journal Week 9
Week 10 cdomin12 Week 10 Class Journal Week 10
Week 11 cdomin12 Week 11 Skinny Genes
Week 12/13 Skinny Genes Quality Assurance Skinny Genes
Week 15 Skinny Genes Deliverables Skinny Genes

Contents

Purpose

To analyze an article for its results, data, and authenticity in terms of how the experiment was conducted and critical analysis.

Outline

1. Ten Biological Terms

2. Outline of Article

1. What is the main result presented in this paper?

The paper showed that there was an impact on gene expression with potassium starvation. It was shown that sulfur metabolism is affected which starts a cascade of events that eventually leads to ammonium accumulation. Ribosome biogenesis and translation decrease expression causing a halt in the cell cycle.

2. What is the importance or significance of this work?

This is important to uncover more about how the role of potassium in the cell and gene expression. This remains mostly unexplored and can reveal pathways in whi

3. What were the limitations in previous studies that led them to perform this work?

Although there have been similar studies conducted before, there is little known about the cellular level in potassium starvation. Due to this, the authors decided to perform this work in order to understand what parts of the cells and function pertain to potassium levels using a newly developed YNB based growth medium and in short term time intervals.

4. How did they treat the yeast cells (what experiment were they doing?)

The yeast cells underwent different amounts of time of potassium starvation in order to access gene expression and cellular function.

5. What strain(s) of yeast did they use? Were the strain(s) haploid or diploid?

Saccharomyces cerevisiae. Haploid or diploid was not mentioned.

6. What media did they grow them in? What temperature? What type of incubator? For how long?

Grown in YNB-based growth medium which has only trace amounts of potassium. Grown at 28 degrees C

7. What controls did they use?

Wild-type BY4741 non-stressed cells.

8. How many replicates did they perform per treatment or timepoint?

Four replicates for each timepoint- 10, 20, 40, 60, and 120 minutes.

9. What method did they use to prepare the RNA, label it and hybridize it to the microarray?

They used wildtype and the experimental and grew them in medium with KCL to 0.8 density. After centrifuging, they then resuspended them into the same media with, and potassium or without. They then took the samples for the microarray at time 10, 20, 40, 60, and 120 minutes.

10. What mathematical/statistical method did they use to analyze the data?

Data Analysis 3.4 software (Bruker Biospion) was used.

11. Are the data publicly available for download? From which web site?

Yes through Society for Applied Microbiology.

12. Briefly state the result shown in each of the figures and tables, not just the ones you are presenting.

Figure 1: This represents induced or expressed gene expression grown on the potassium free medium at the relative time points. These measurements were made through RT-PCR assays. There is a pretty even amount of induced and repressed genes

Table 1: This table shows the transcriptional changes at minutes 10, 20, 40, 60, and 120. It measured induction/repression also measured through RT-PCR assays. No trend to show here.

Table 2: Where the strains were taken from in regard to work from other papers.

Figure 2: This pertains to the cellular function of sulfur amino acid metabolism. It has three parts to it. A shows the mechanism of how potassium starvation effects metabolism. B with the X axis representing mRNA change and Y axis being different genes shows the mRNA change in the different time intervals. C (x axis as time min and Y as intracellular levels of methionine and cysteine) shows.

Figure 3: One graph (x axis- time in minutes and y axis changes in expression) shows different genes and their change in expression over time. The other graph (x axis- time in minutes and y axis glutathione levels) shows glutathione levels over time. Both of these show that there is no general stress response to potassium starvation.

Figure 4: (A) This demonstrates the effect of methylglyoxal production as well as trehalose metabolism from potassium starvation. (B) Two graphs (x axis- time in min and y axis- changes in expression fold) shows the change in expression specifically for genes related to trehalose metabolism and methylglyoxal. Measurements were made through the microarray data. (C) A graph (x axis- time in min and y axis- methylglyoxal concentration) showing MG levels remaining higher after 20 minutes potentially linked to genes induced in trehalose metabolism.

Figure 5: (A) Graph (x axis- time in min and y axis- change in expression) shows genes seen in mitochondrial retrograde response. Measurements made through microarray data. DIT2 and DLD2 response done through measurements of microarray data. This shows electrophoresis data for it. (B) mitochondria morphology picture with mutant showing different morphology. Measured through GFP microscopy. (C) Graph (x axis- time min and y axis- ammonium concentration) shows concentration increase after being put in potassium free medium. (D) Graph (x axis- time in min and y axis- ammonium concentration) showing mutated Trk1 or Trk2 did not have more concentration. Measured through samples taken with transformed plasmids.

Figure 6: (A) Graph (x axis- time in min and y axis- changes in expression) shows different genes during potassium starvation. Measured through microarray experiments. (B) mRNA abundance shows of different cyclins measured through RT-PCR. (C) Demonstrates the cyclin data showing that they are almost undetectable. Measured through immunoblot analysis.

Figure 7: (A) Graph (x axis- time in min and y axis- change in expression) of genes that are involved in septin ring formation. Measured through microarray data. (B) Pictures showing + or – potassium after different amounts of time. (C) Measured through immunoblot analysis shows levels of Cdc11 protein. (D) wildtype measured through immunofluorescence with and without KCl.

13. How does this work compare with previous studies?

They wanted to specifically examine cellular functions of potassium related to time points in short-term starvation. They also were able to use a newly developed YNB based growth medium which had not been used in previous studies before. This allowed them to grow strains that were not exposed to any potassium.

14. What are the important implications of this work?

To learn more about the cellular function of potassium, why it is important and how it can be affected. It is clear that there are connections to other parts of cellular function that cannot perform or do as it should without potassium in the cell.

15. What future directions should the authors take?

They should look more closely at the pathways that were affected and the relative transcription factors. They were able to more closely characterize more of cellular functions effected by potassium; however, now they are more aware of which are effected they can dive deeper into why and how it plays a role.

16. Give a critical evaluation of how well you think the authors supported their conclusions with the data they showed. Are there any major flaws to the paper?

They had a lot of data to support what they were concluding. They were able to have the evidence for everything conclusion that they made. Saying that, I found the paper to be confusing with the ample amounts of results that they had and I did not know exactly why they performed certain experiments or could not figure out the methodology as to how it related to their question.

Annotated Bibliography

    1. Use a keyword search for the first three databases/tools and answer the following:
      • PubMed
        • What original keyword(s) did you use? How many results did you get?
        • Which terms in which combinations were most useful to narrow down the search? How many results did you get after narrowing the search?
      • Google Scholar
        • What original keyword(s) did you use? How many results did you get?
        • Which terms in which combinations were most useful to narrow down the search? How many results did you get after narrowing the search?
      • Web of Science
        • What original keyword(s) did you use? How many results did you get?
        • Which terms in which combinations were most useful to narrow down the search? How many results did you get after narrowing the search?
    2. Use the advanced search functions for each of these three databases/tools and answer the following:
      • PubMed
        • Which advanced search functions were most useful to narrow down the search? How many results did you get?
      • Google Scholar
        • Which advanced search functions were most useful to narrow down the search? How many results did you get?
      • Web of Science
        • Which advanced search functions were most useful to narrow down the search? How many results did you get?
      • Perform a prospective search on your article in the Web of Science and answer the following:
        • How many articles does this article cite?
        • How many articles cite this article?

Bibliographic Entries

  1. Schade, B., Jansen, G., Whiteway, M., Entian, K.D., & Thomas, D.Y. (2004). Cold Adaptation in Budding Yeast. Molecular Biology of the Cell, 15, 5492-5502. doi: 10.1091/mbc.E04-03-0167
  2. PubMed Abstract: http://www.ncbi.nlm.nih.gov/pubmed/15483057
  3. PubMed Central: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC532028/
  4. Publisher Full Text (HTML): http://www.molbiolcell.org/content/15/12/5492.long
  5. Publisher Full Text (PDF): http://www.molbiolcell.org/content/15/12/5492.full.pdf+html
  6. Copyright: 2004 by the American Society for Cell Biology (information found on PDF version of article);
  7. Article is not Open Access, but is freely available 2 months after publication, published in the United States, LMU did not pay for the article
  8. Availability: in print and online
  9. Publisher: American Society for Cell Biology (scientific society), non-profit, not a member of the OAPA
  10. The journal begain in 1989
  11. Peer-reviewed articles: yest
  12. Editorial board.
  13. Impact factor: 3.512 (2017)
  14. Primary research article
  15. Data are not available (dead hyperlink)


  1. Schade, B., Jansen, G., Whiteway, M., Entian, K.D., & Thomas, D.Y. (2004). Cold Adaptation in Budding Yeast. Molecular Biology of the Cell, 15, 5492-5502. doi: 10.1091/mbc.E04-03-0167
  2. PubMed Abstract: http://www.ncbi.nlm.nih.gov/pubmed/15483057
  3. PubMed Central: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC532028/
  4. Publisher Full Text (HTML): http://www.molbiolcell.org/content/15/12/5492.long
  5. Publisher Full Text (PDF): http://www.molbiolcell.org/content/15/12/5492.full.pdf+html
  6. Copyright: 2004 by the American Society for Cell Biology (information found on PDF version of article);
  7. Article is not Open Access, but is freely available 2 months after publication, published in the United States, LMU did not pay for the article
  8. Availability: in print and online
  9. Publisher: American Society for Cell Biology (scientific society), non-profit, not a member of the OAPA
  10. The journal begain in 1989
  11. Peer-reviewed articles: yest
  12. Editorial board.
  13. Impact factor: 3.512 (2017)
  14. Primary research article
  15. Data are not available (dead hyperlink)

Reflect

1.What impact does the choice of keywords have on your results?

2.What are the advantages and disadvantages of each of the three databases (Pubmed, Google Scholar, Web of Science)?

  • Google Scholar: Disadvantages include limited capability of narrowing down search and not including DOI number in citations. Advantages include formatted citations and a familiar interface that is user friendly.
  • Pubmed: Disadvantages include keywords allowing for specific filters that can limit what is presented as a result. Advantages include a more detailed query for narrowing down searches.
  • Web of Science: Disadvantages include less up to date results and less relevance in results. Advantages include all articles related to sciences which allows for more diverse results and a university resource. They also have a citation network that allows for connections to all papers referenced.

Conclusion

While working through the outline, this paper had a lot of results and figures to represent the data. It was a bit confusing and misleading at times, where I had to reread the article multiple times. I felt that this article could have been explained better in a manner that led better into the results and figures. It was difficult to work with, but it did have findings that seemed promising in the field of research.

Acknowledgments

1. I worked with User:ymesfin, User:Jcowan4, User:msamdars, and User:dramir36 for this assignment.

2."Except for what is noted above, this individual journal entry was completed by me and not copied from another source." Cdomin12 (talk) 19:53, 11 November 2019 (PST)

References

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