Expression Analysis of the Sphingolipid Metabolism

1
Expression Analysis of the Sphingolipid
Metabolism
?
Download GenMAPP v2.1
?
Gene Extraction
???
Data Formatting
?V
Download Pathways
V
V?
Pathway Modification
Data Application
Branch Addition
Gene Addition
2
I. GenMAPP v2.1
GenMAPP v2.1, a Windows operating
program, provides a technique for conducting a
genomic analysis through the
visualization of gene expression data within a
metabolic pathway. Expression data derived
from microarray and other similar genomic
experiments can be imported and recognized by
GenMAPP using multiple gene
identifiers such as Entrez Gene, Ensembl, and
Affy ID's. This program allows for
the genes within a biological pathway, such as
the sphingolipid metabolism, to be color-coded
based on a profile established by
the user. In addition, tools provided within
GenMAPP allow such metabolic pathways
to be created or edited by the user. The
figure shown below demonstrates an example of
what one would obtain after
importing expression data into GenMAPP.
www.genmapp.org
3
II. How to Extract Sphingolipid Related Genes
If desired genes have been previously
extracted, proceed to step ???.
Perl program instructions
The perl script accepts a text file with the list
of gene symbols or the tag IDs of interest. It
opens the microarray file with normalized gene
expression values and searches for rows that
match with the gene symbol or Tag IDs .The
matched rows are copied to a new test file that
may be opened using excel and further data
analysis can be performed.
Program Name Affyfilter.pl
Function Script for filtering gene expression
values of interest from Normalized gene
expression file
Requirement Perl 5.8 must be down loaded and
installed on the windows system from
www.activestate.com without charge. Mac s have
built in perl in their X11 command line.
USE Make sure to have program file
(affyfilter.pl) , genelist file and normalized
microarray file in the same folder where perl is
installed.
At the command line type C\ perl gt
perl affyfilter.pl microarray.txt genelist.txt
gt output.txt
The results of the filtering will be stored in a
output file within the same folder (Note that the
names of the file can are user defined)
Example of a text file with gene list
Example of excel file with normalized gene
expression values
4
III. How to Format Data in GenMAPP
GenMAPP recognizes data to be imported in a
specific format as illustrated below. Column
headings GeneID, System Code, and a user given
data heading (e.g. Control), must be included for
proper identification. The system code is a code
provided by GenMAPP corresponding to the gene
database used for identifying the GeneID. The
data sheet should be saved as a text document
(.txt), tab-delimited file (.tab), or
comma-separated values (.csv).
Gene Identification Code Corresponding
Data 1
to Gene Database
GeneID System Code Control
54218 L 1.43
14421 L 0.51
14421 L 0.47
12091 L 2.07
- .txt, .tab, or .csv file
System Codes EntrezGene L MGI
M Ensemble En UniProt S Unigene
U Affy X
5
IV. Download Sphingolipid Pathways
Download Sphingolipid pathways (.mapp file) from
www.sphingomap.com ? Sphingo-GeneMap and open
within GenMAPP. Available Pathways
Include De novo
Lacto Ganglio
Neolacto IsoGlobo
Globo
6
V. How to Modify Provided Sphingolipid Pathways
- Gene Addition

1. Select species specific database, See Gene
   Database Selection
2. Click on gene box located in the tool bar of the
   draftboard
3. Right click gene to open Gene Finder
4. Enter the gene ID and the corresponding gene ID
   system
5. Within Gene Label enter gene name

7
V. How to Modify Provided Sphingolipid Pathways
- Branch Addition
8
VI. How to Apply Data to Pathway Using GenMAPP
1. Gene Database Selection

1. Data ? Download Data from GenMAPP.org ?
   expand Gene Database and select species used
2. Within Database File Name, select database to
   be used and click start
3. Return to draftboard
4. Data ? Choose Gene Database ? select
   database downloaded

9
VI. How to Apply Data to Pathway Using GenMAPP
2. Creating a Color-Criteria

• Data ? Expression Dataset Manager
• Expression Datasets ? New Dataset ? select
  saved data file
• Color Sets ? New
• Using Boolean expression values provided,
  establish color-criteria based on data
• Save Expression File and exit Expression Dataset
  Manager
• Values in GenMAPP represent
  Fold Changes

10
Sphingolipid Metabolism
Detailed Instructions, list of additional
functions available, and downloads are
available at http//www.genmapp.org/