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miRvestigator Framework Tutorial

Important Note: At any point along the process help information are embedded in the web page as [+] icons which once clicked will drop down information about the task being performed. The help can easily be retracted by clicking on the [-].

 
  1. Enter a list of Entrez gene IDs corresponding to co-expressed genes
  2.  

     
    1. If you want to use your own data:
    2.  
      1. Identify putatively co-regulated genes (e.g. co-expressed genes)
      2.  
      3. Convert the gene identifiers to either Entrez gene, Ensembl gene, RefSeq transcript identifiers or official gene symbols using the DAVID bioinformatics resource gene ID conversion tool (Note: DAVID has great a help page for ID conversion.)
      4.  
      5. Choose the species for your data
      6.  
      7. Decide whether to include viral miRNAs
      8.  
      9. Choose the gene identifier type for your data
      10.  
      11. Name your gene list for easier identification of analysis conducted
      12.  
      13. Input the genes into the miRvestigator interface
       
    3. Or click one of the sample data buttons to load sample data (Note: hsa-miR-9 is small so it runs quite fast for a tutorial.)
     
     
  3. Set parameters for the run (Note: For most cases the default parameters will suffice.)
  4.  

     
    1. Set Weeder parameters
      1.  
      2. Choose whether you want to search for 6bp and/or 8bp motifs
      3.  
      4. Choose background model for Weeder to compare enriched motifs against
      5.  
      6. Choose target site quality threshold to filter predicted binding sites
       
    2. Set the miRvestigator parameters
      1.  
      2. Choose which seed models you wish to investigate
      3.  
      4. Choose whether you want to search for wobble base-pairing or not
        •  
        • If you do want to search for wobble base-pairing then choose the minimum G or U frequency
     
     
  5. Submit the job
  6.  

     
    1. Choose the number of miRNAs to return
    2.  
    3. Choose whether you want to be notified by e-mail or not (Note: Giving an e-mail address is completely optional, although it can be nice to have an e-mail with a link to the results.)
    4.  
    5. Click the submit button
     
     
  7. Watch as the job runs (Note: You can bookmark this page to come back for your results.)
  8.  

     
    1. Status events as miRvestigator runs:
      1.  
      2. Extracting Regulatory Sequences
      3.  
      4. Running Weeder
      5.  
      6. Running miRvestigator HMM
      7.  
      8. Compiling Results
       
    2. Estimated run time for the submitted job
    3.  
    4. Submitted parameters are shown below the status information
     
     
  9. Results for the run are returned (Note: You can bookmark this page to come back for your results.)
  10.  

     
    1. To see what parameters were used for the run click on the [+] to expand the Submission Parameters table
    2.  
    3. The Summary of Results table has an entry for each motif identified in the 3 UTRs for the input sequences
      1.  
      2. Click on the Motif link to be brought to the Motif Complementary miRNAs table. You can also download the table as a CSV (comma separated values) formatted file for import into a spreadsheet application like Microsoft Excel. (Note: The help [+] for this table describes the columns in detail.)
      3.  

         
           
        • Good quality complementarity between motif and miRNA have Complementarity P-Values equal to 1.5e-05 or 6.1e-05 for perfect 8mer or 7mer complementarity, respectively
        •  
        • The Complementary Base-Pairing column gives a textual representation of the complementary base-pairing
         
      4. Click on the % of Input Sequences with Site to be brought to the Motif Sites table. You can also download the table as a CSV (comma separated values) formatted file for import into a spreadsheet application like Microsoft Excel. (Note: The help [+] for this table describes the columns in detail.)
      5.  

         
           
        • Good quality motif sites have higher similarity and are colored red
        •  
        • High quality sites from this table would be excellent candidates for experimental studies
     
     
  11. Returning to Access Results at a Later Date
    1.  
    2. Bookmark Status page
    3.  
    4. Bookmark Results page
    5.  
    6. Add e-mail address to run parameters receive e-mail with link to results (optional)

Need help? Please contact cplaisier(at)systemsbiology.org or cbare(at)systemsbiology.org if you have any questions, comments or concerns.
Developed at the Institute for Systems Biology in the Baliga Lab.