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  1. IGB
  2. IGBF-3472

Improve information on interaction term in markdown

        Details

        • Type: Task
        • Status: Closed (View Workflow)
        • Priority: Major
        • Resolution: Done
        • Affects Version/s: None
        • Fix Version/s: None
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          Description

          File: FindTreatmentEffectAcrossGenotypes-DESeq2.Rmd

          Check to see if the analysis and the interaction term is explained well and provides enough sanity checking to confirm no significance.
          Use bar plot shiny app to show analysis.

          During meeting with Muday Lab, we found a gene that we thought ought to have come up in a test of the interaction term: Solyc02g063355.1

          The SL5 version of that gene appears in the top row of the attached heat map file 2023-10-19-meeting-heatmap-visualization.png.
          The heat map shows log2(average(cpm(A.28/V.28))) for treatment durations indicated on the x axis of the plot.

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              Mdavis4290 Molly Davis added a comment - - edited

              Confirming the Analysis with bar plots of gene expression:

              • Changed log-fold change threshold to 0.5 and only got 1 DE gene at just 30 min. Changed to 0.1 and got 5 DE genes at just 30 min. Changed to 0.05 and still only got the exact same 5 DE genes at 30min from before. Change to 0 fold change threshold and got 6 DE genes at 30 min. These genes are:
              1. Solyc09g092690.3
              2. Solyc04g011440.4
              3. Solyc06g076560.2
              4. Solyc01g102960.3
              5. Solyc03g117630.1
              6. Solyc03g120090.1
              • Reviewed the 6 DE genes at 30 min in the expression barplot Dr. Loraine made to confirm analysis (https://ann-loraine-gmail.shinyapps.io/72_F3H_PollenTube/) seen above. These genes meet the statistical design of being DE across genotype VF36 plus ARE and the temperatures 28C and 34C. In other words, we are answering the question: is the condition effect different across genotypes?
              • Wrote in more detail of what was used and why in the markdown.
              Show
              Mdavis4290 Molly Davis added a comment - - edited Confirming the Analysis with bar plots of gene expression : Changed log-fold change threshold to 0.5 and only got 1 DE gene at just 30 min. Changed to 0.1 and got 5 DE genes at just 30 min. Changed to 0.05 and still only got the exact same 5 DE genes at 30min from before. Change to 0 fold change threshold and got 6 DE genes at 30 min. These genes are: Solyc09g092690.3 Solyc04g011440.4 Solyc06g076560.2 Solyc01g102960.3 Solyc03g117630.1 Solyc03g120090.1 Reviewed the 6 DE genes at 30 min in the expression barplot Dr. Loraine made to confirm analysis ( https://ann-loraine-gmail.shinyapps.io/72_F3H_PollenTube/ ) seen above. These genes meet the statistical design of being DE across genotype VF36 plus ARE and the temperatures 28C and 34C. In other words, we are answering the question: is the condition effect different across genotypes? Wrote in more detail of what was used and why in the markdown.
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              Mdavis4290 Molly Davis added a comment - - edited

              Branch: https://bitbucket.org/mdavis4290/molly3-flavonoid-rnaseq/branch/IGBF-3472

              Reviewer:

              • Please pull changes from FindTreatmentEffectAcrossGenotypes-DESeq2.Rmd markdown branch and make sure everything runs properly with knit
              • Review writing and comprehension of the analysis
              • Is the interaction term and log2foldchange explained properly
              • Are the output files present in results folder (MvW-temp-[assembly].xlsx) and structurally make sense
              • Do the barplots in the previous comment of the 6 DE genes support the analysis? If so then they can also be added to the Flavonoid repo. Another ticket could be made for this.
              Show
              Mdavis4290 Molly Davis added a comment - - edited Branch : https://bitbucket.org/mdavis4290/molly3-flavonoid-rnaseq/branch/IGBF-3472 Reviewer: Please pull changes from FindTreatmentEffectAcrossGenotypes-DESeq2.Rmd markdown branch and make sure everything runs properly with knit Review writing and comprehension of the analysis Is the interaction term and log2foldchange explained properly Are the output files present in results folder (MvW-temp- [assembly] .xlsx) and structurally make sense Do the barplots in the previous comment of the 6 DE genes support the analysis? If so then they can also be added to the Flavonoid repo. Another ticket could be made for this.
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              robofjoy Robert Reid added a comment - - edited

              I have pulled down the code.

              Running this on console because it asked me to.
              tinytex::install_tinytex()

              Show
              robofjoy Robert Reid added a comment - - edited I have pulled down the code. Running this on console because it asked me to. tinytex::install_tinytex()
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              robofjoy Robert Reid added a comment -

              I had to re-install EdgeR package for some reason.
              But once I did and installing the tinytex library, everything knitted !

              So now I have a PDF.

              Reading through the PDF now to see what is there....

              Show
              robofjoy Robert Reid added a comment - I had to re-install EdgeR package for some reason. But once I did and installing the tinytex library, everything knitted ! So now I have a PDF. Reading through the PDF now to see what is there....
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              robofjoy Robert Reid added a comment -

              Ran the code in RStudio.

              Got the 2 xlsx files (SL4 and SL5).
              They look to have 4 tabs, one for each time point.

              And look as one would expect.

              As expected, those p-values leave us with nothing!
              That does seem to not coincide with the bar plots that we see in the comments above. So confusing.

              Show
              robofjoy Robert Reid added a comment - Ran the code in RStudio. Got the 2 xlsx files (SL4 and SL5). They look to have 4 tabs, one for each time point. And look as one would expect. As expected, those p-values leave us with nothing! That does seem to not coincide with the bar plots that we see in the comments above. So confusing.
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              Mdavis4290 Molly Davis added a comment - - edited

              There should be 6 DE genes when the threshold for log2foldchange is 0. The code from the branch in this ticket should have a threshold of 0 and the 6 significant genes are only present at 30 minutes. The code is at the very top of the markdown that indicates the threshold. Do you see this occurrence? It is also stated in the knitted pdf how many DE genes are present at each time chunk. The 6 genes I found to be significant are the 6 genes in the barplots in the above comment. Thanks!
              [~RobertReid]

              Show
              Mdavis4290 Molly Davis added a comment - - edited There should be 6 DE genes when the threshold for log2foldchange is 0. The code from the branch in this ticket should have a threshold of 0 and the 6 significant genes are only present at 30 minutes. The code is at the very top of the markdown that indicates the threshold. Do you see this occurrence? It is also stated in the knitted pdf how many DE genes are present at each time chunk. The 6 genes I found to be significant are the 6 genes in the barplots in the above comment. Thanks! [~RobertReid]
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              robofjoy Robert Reid added a comment - - edited

              Confirmed !!

              I see the 6 genes below the P-adjusted cutoff at 30 minutes (SL4) once the LFC threshold is set to 0.

              Are there any other pieces here that need review?

              Show
              robofjoy Robert Reid added a comment - - edited Confirmed !! I see the 6 genes below the P-adjusted cutoff at 30 minutes (SL4) once the LFC threshold is set to 0. Are there any other pieces here that need review?
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              Mdavis4290 Molly Davis added a comment -

              That is all that needs to be reviewed. Thank you!

              Creating a PR!

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              Mdavis4290 Molly Davis added a comment - That is all that needs to be reviewed. Thank you! Creating a PR!
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              Mdavis4290 Molly Davis added a comment -

              Pull Request: https://bitbucket.org/hotpollen/flavonoid-rnaseq/pull-requests/29

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              Mdavis4290 Molly Davis added a comment - Pull Request : https://bitbucket.org/hotpollen/flavonoid-rnaseq/pull-requests/29
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              Mdavis4290 Molly Davis added a comment -

              Testing:

              • Had no issues knitting the markdown.
              • Results files are saved and organized properly.
              • Comprehension & grammar are fine. Might need to improve further later on.

              Moving to done!

              Show
              Mdavis4290 Molly Davis added a comment - Testing : Had no issues knitting the markdown. Results files are saved and organized properly. Comprehension & grammar are fine. Might need to improve further later on. Moving to done!

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                • Assignee:
                  Mdavis4290 Molly Davis
                  Reporter:
                  Mdavis4290 Molly Davis
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                    Updated:
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