GV Exercise.4

Find differentially expressed genes and perform functional enrichment

[ Main_Page | Genevestigator_training | Analyze_public_microarray_data_using_Genevestigator | GV Exercise.3 |
| GV Exercise.5 ]
last edit: October 31, 2014

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Aim

Use the Condition Search Tools to identify Samples, Anatomy, Neoplasm (when applies), Perturbations, or Deverlopment annotations that are significantly linked to a list of probes.

Find perturbations where metacaspases are differentially expressed

• select Ath.all as Sample Selection on the top-left of the window and metacaspases as Gene selection at the bottom-left.
• open the Condition Search Perturbations tool
• change the data display to heatmap-list; if using -tree, choose expand all

We get a nice but very long heatmap with many lines showing little if any differential expression

Note the two popups on the right, it allows filtering for more pronounced effects and get shorter heatmaps

Use |2| fold change and pval <0.05 to restrict the heatmap to most significant treatments for the leftmost probe (ATMCPB1 / LOL3)

• How many treatments are kept?
• Which other metacaspase(s) often show DE (UR and DR) for the same treatments?

Create a new Sample Selection out of Ath.all based on the filtered perturbations

create a 'new' sample selection after filtering and give the selection a good name

Name your new Sample Selection: ATMCPB1_filtered-pert, it will be used later and was saved as part of ex4.gv4 (see bottom of this page).

Change now to the Anatomy tab and click on probe headers to resort the lines

• Set the Sample selection back to Ath.all
• Which tissues do show the highest (lowest) potential expression for the leftmost probe? Are other probes sharing a similar pattern?

Change now to the Development tab and look at the plant stages for metacaspase expression levels

• Alternate between Scatterpot and Heatmap display to estimate which view fits you best.
• export the results to an excel formatted Development-metacaspases.xls file on your laptop then open this file using installed software (eg. libre-office)

metacaspases during development (scatter or heatmap)

Apply Gene Ontology Enrichment to the list of caspases with PLAZA

Copy the list of TAIR IDs from the exported file or from below

list of TAIR10 IDs for metacaspases

# Ath metacaspase (TAIR10)
AT1G02170
AT5G64240
AT1G79310
AT1G79320
AT1G79330
AT5G04200
AT4G25110
AT1G79340
AT1G16420

Register to gain access to PLAZA (dicot fro Ath) then log-in and navigate to Analyze' // 'workbench (http://bioinformatics.psb.ugent.be/plaza/versions/plaza_v3_dicots/workbench/logon)

Create a new experiment and paste the metacaspase list in the provided text box, save the result with a meaningful title

In the section dedicated to Viewing, pick .. the GO enrichment

GO enrichment metacaspases

There is OBVIOUSLY MORE than just GO in Plaza ...

Perform GSEA with the Plant GSEA web-portal

Copy the list of TAIR IDs from the exported file or from below

list of TAIR10 IDs for metacaspases

# Ath metacaspase (TAIR10)
AT1G02170
AT5G64240
AT1G79310
AT1G79320
AT1G79330
AT5G04200
AT4G25110
AT1G79340
AT1G16420

Navigate to Plant GSEA (http://structuralbiology.cau.edu.cn/PlantGSEA/) Paste the metacaspase list in the Analysis page and select what applies (run the tool)

Inspect the resulting tables (not reproduced here) and the bottom matrix.

GO enrichment metacaspases (2013: left, 2014: right)

See how much public database content can vary upon time (one year) !!

Download the exercise files

Try it by yourself before expanding on the right!

• download Ath_metacaspases_TAIR.txt and open it with your default worksheet application file link
• download Development-metacaspases.xls and open it with your default worksheet application file link
• download ex4.gv4 and open it from within genevestigator File Load Workspace file link

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References:

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| GV Exercise.5 ]