Richard Brous Week 9

Richard Brous

• Week 9 Assignment
• RAB_week9_archive.zip

Electronic Lab Notebook - Analysis of Vibrio Cholerae microarray data project for Week 9

Getting Started

• Partnered with Zeb (older db)
• I have selected the newest Vc-Std_External10201022 gene database
  • Extracted to Desktop

Staring GenMAPP and getting organized

• Launched GenMAPP (thank god its not looking for update server!!! =D)
• Ensure correct gene database is loaded Vc-Std_External10201022
  • If not, load the correct one
    • Data > Choose Gene Database menu item to select the Gene Database you need
• Select Data menu then Expression Dataset manager
  • Select new dataset which is the tab deliminated text file formatted for GenMAPP made during week 8.
  • Since the tab-deliminated file contains only data DO NOT select any column to exclude from Data Type Specification window
  • Let the Expression dataset manager convert the data
    • This may take a while so don't think system hung up... only minimal screen feedback
      • Complete conversion yields the data active in the Expression Dataset Manager window AND creates a conversion file named *.gex placed in the same location as where your text-deliminated sourcefile was.
    • Errors (almost certainly) occured where the Expression Dataset manager could not convert 1 or more lines of data
      • An exception file is created which contains all the raw data with the addition of a column called "~Error~"
        • Errors are either messages or if the program finds no errors: a single space character
      • I specifically found 121 errors of 5221 records
      • Zeb should get more errors since his gene database is older.
  • Customize the new Expression Dataset by creating new Color Sets
    • Color Sets contain the GenMAPP instructions for displaying data from an Expression Dataset on MAPPS.
      • Create a color set by filling in the fields:
        • Name for the Color Set: pathogenic_vs_lab
        • gene value: Avg_LogFC_all
        • Criteria that determine how a gene object is colored on the MAPP
          • increased gene expression (red)
            • [Avg_LogFC_all] > 0.25 AND [Pvalue] < 0.05
          • decreased gene expression (green)
            • [Avg_LogFC_all] < -0.25 AND [Pvalue] < 0.05
          • Expressions are equivalent to queries performed in PostgreSQL
    • After completing a new criterion, add the criterion entry (label, criterion, and color) to the Criteria List by clicking the Add button
    • 2 criterion were created: "Increased" will be [Avg_LogFC_all] > 0.25 AND [Pvalue] < 0.05 and "Decreased will be [Avg_LogFC_all] < -0.25 AND [Pvalue] < 0.05
    • Can always add more criterion by following previous steps
    • Save the entire Expression Dataset by selecting Save from the Expression Dataset menu
  • Exit Expression Dataset Manager to view the Color Sets on a MAPP and then close it
• Keep your *.gex file safe prior to wiki upload (save to usb drive or email it to yourself)

MAPPFinder Procedure

• Zeb and I were in front row selected: INCREASED
• Launch MAPPFinder program or launch from within GenMAPP select Tools -> MAPPFinder
• Ensure the correct Gene Database is loaded!!!
  • If not choose File -> Choose Gene Database and select the correct one.
    • Located in: C:\GenMAPP 2 Data\Gene Databases\
• Press "Calculate New Results" button
  • Select your *.gex file you created previously.
    • RAB_10_23_2010_Merrell_Compiled_Raw_Data_Vibrio.gex
  • Click OK
  • Choose the Color Set and Criteria with which to filter the data
    • Chose Increased since that is what we are assigned
    • Check Gene Ontology and Calculate p values
    • Click the "Browse" button and create a meaningful filename for the results
    • Click "Run MAPPFinder".
      • The analysis will take several minutes.
        • It may look like the computer is stalled; but the hourglass should be on the screen indicating its working
• When the results have been calculated, a Gene Ontology browser will open showing your results
  • All of the Gene Ontology terms that have at least 3 genes measured and a p value of less than 0.05 will be highlighted yellow.
    • A term with a p value less than 0.05 is considered a "significant" result.

Evaluating MAPPFinder results

• List most significant Gene Ontology terms
  • Click on "Show Ranked List" from Menu Bar
• Top 10 Gene Ontology terms:
  • branched chain family amino acid metabolic process
  • branched chain family amino acid biosynthetic process
  • IMP biosynthetic process
  • IMP metabolic process
  • arginine metabolic process
  • cellular nitrogen compound biosynthetic process
  • leucine biosynthetic process
  • leucine metabolic process

1. amine biosynthetic process
2. arginine biosynthetic process