Notes installing the SBEAMS - Microarray_Affy module Background) ISB developed the microarray module to initially deal with organizing and analyzing 2-color data from arrays printed in-house. Starting around July 2004, development started to extended the Microarray module to organize and analyze the Affymetrix Expression array data. The install instructions are for setting up just the Affy portion of the module. You may notice that some tables and links are not utilized by the Affy portion of the Microarray module but are still visible since both the 2-color and Affy array infrastructure were built out within the same sbeams module. ------------------------------------------------------------------------------- 1) Software and module Dependencies You must first install the SBEAMS Core. See the separate installation notes (sbeams.installnotes) on how to accomplish that. You must also install the BioLink module; follow the installation instructions provided with that module first (BioLink.installnotes). The following Perl Modules often not found on a standard UNIX/Linux setup are required to successfully use SBEAMS - Microarray_Affy (in addition to the dependencies for the SBEAMS Core). XML::LibXML; Tie::IxHash; XML::Writer Archive::Zip Statistics::R [Optional] Not used in production code. Only used in a test script Make_R_graph.cgi to demonstrate making a graph in using R and Perl --------------- The following non-Perl software is required: Netpbm (http://netpbm.sourceforge.net) R -Version 2.0 or greater R -Bioconductor libraries, see below. library(affy) library(gcrma) library(vsn) library(Biobase) library(multtest) library(annaffy) library(webbioc) library(siggenes) You can install all these libraries at once from within your R (after you've installed it) by running these 2 commands: source("http://www.bioconductor.org/getBioC.R") getBioC() #XSLT transformation engine {semi-optional} Only need to view the help pages Ehttp://xmlsoft.org/XSLT/ xsltproc ------------------------------------------------------------------------------- 2) Installation Location SBEAMS is designed to live entirely in the "htdocs" area of your Apache web server. For the remainder of this installation, it will be assumed that your installation is configured as follows; compensate for your specific setup: servername: db DocumentRoot: /local/www/html Primary location: directly located in DocumentRoot, /local/www/html/sbeams --> http://db/sbeams/ Development location: In a dev1 tree starting in the DocumentRoot, /local/www/html/dev1/sbeams -> http://db/dev1/sbeams/ All modules live in the same area and should be unpacked into the main SBEAMS area. # Set $SBEAMS enviroment variable (tcsh syntax): setenv SBEAMS /local/www/html/dev1/sbeams # bash syntax: # export SBEAMS=/local/www/html/dev1/sbeams ------------------------------------------------------------------------------- 3) Create and populate the database It is assumed that you have already created and tested your SBEAMS Core database. You may either create a separate database for the Microarray_Affy database or you can put everything in the same database. Note that some database engines (rare now) may not permit cross-database queries in which case your may NOT use separate databases. If you do use separate databases, you may not be able to enforce referential integrity between tables in the different databases. This may or may not be a significant concern. - If you decide on a separate database, create it and within it, create users "sbeams" and "sbeamsro" as a read/write account and a read-only account, respectively, as done for the Core. - Generate the appropriate schema for your type(s) of database as follows: cd $SBEAMS/lib/scripts/Core set dbtype=mssql #### Set to your RDBMS type: mssql, mysql, etc. ./generate_schema.pl \ --table_prop ../../conf/Microarray/Microarray_table_property.txt \ --table_col ../../conf/Microarray/Microarray_table_column.txt \ --schema_file ../../sql/Microarray/Microarray --module Microarray \ --destination_type $dbtype --suppress - Verify that the SQL CREATE and DROP statements have been correctly generated in $SBEAMS/lib/sql/Microarray/ - Execute the statements to create and populate the database with some bare bones data and indexes (for faster loading and querying). This uses the generic SQL execution script runsql.pl. Note that several of the commands below use the -i flag, this allows installation to continue in case there are errors. Currently there are such errors, so you may well see some error output from the script. SQL Server Example: To CREATE and POPULATE: ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATETABLES.mssql ./runsql.pl -u sbeamsadmin -s $SBEAMS/lib/sql/Microarray/Microarray_POPULATE.sql ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATECONSTRAINTS.mssql To CREATE indexes: ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATEINDEXES.sql To CREATE Manual Constraints (Constraints that could not be auto-generated) ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_ADD_MANUAL_CONSTRAINTS.mssql ##### MySQL ############### # # Copy: # Microarray_ADD_MANUAL_CONSTRAINTS.mssql to Microarray_ADD_MANUAL_CONSTRAINTS.mysql # # Edit Microarray_ADD_MANUAL_CONSTRAINTS.mysql to remove dbo. from beginning # of table name on each line, and add a semicolon at the end of each line # # Edit Microarray_CREATEINDEXES.sql to remove dbo. from the beginning of each # table name and remove all comment lines (which begin with '--') # # Execute: # ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATETABLES.mysql --delimiter semicolon # ./runsql.pl -u sbeamsadmin -s $SBEAMS/lib/sql/Microarray/Microarray_POPULATE.sql --delimiter semicolon # ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATECONSTRAINTS.mysql --delimiter semicolon # ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_CREATEINDEXES.sql --delimiter semicolon # ./runsql.pl -u sbeamsadmin -i -s $SBEAMS/lib/sql/Microarray/Microarray_ADD_MANUAL_CONSTRAINTS.mysql --delimiter semicolon # ########################### Note that the Microarray_POPULATE.mssql is not auto-generated and should probably work for all flavors of database Examples for table creation for other database flavors can be found in the Core installation notes and will not be repeated here. Notes: A few warnings or errors might be seen while creating the tables and constraints. The ones below should be considered normal DBD::Sybase::db do failed: Server message number=2714 severity=16 state=6 line=3 server=TITAN text=There is already an object named 'biosequence_set' in the database. at /var/www/html/sbeams/lib/scripts/Core/runsql.pl line 113, line 866. ------------------------------------------------------------------------------- 3-1) Data storage locations and naming conventions ISB Affymetrix Core facility name it's files with the following convention All Files generated by the core facility CEL, CHP, XML, RPT files all use the common root name YYYYMMDD_DD_. YYYYMMDD = Date array was scanned DD = Unique Scan Number per Day ie 01, 02, 03.... Sample_name = User Sample Name, No Spaces or strange characters Data Location. All the data is located within one main directory which is subdivided into folders named YYYYMM - For external datasets we have a folder "external" within the main data repository and individual experiments are separated into a user defined folder A cron job can be setup (see below) to keep monitoring the default directory for any new files to be uploaded ######################### For initial setup make the following folders # These should already exist from the installation of the Core module mkdir $SBEAMS/tmp $SBEAMS/var # These will have to be created. mkdir $SBEAMS/tmp/Microarray \ $SBEAMS/tmp/Microarray/AFFY_ANNO_LOGS \ $SBEAMS/tmp/Microarray/R_CHP_RUNS \ $SBEAMS/tmp/Microarray/GetExpression \ $SBEAMS/tmp/Microarray/GetExpression/jws \ $SBEAMS/var/Microarray \ $SBEAMS/var/Microarray/Affy_data \ $SBEAMS/var/Microarray/Affy_data/probe_data \ $SBEAMS/var/Microarray/Affy_data/probe_data/external \ $SBEAMS/var/Microarray/Affy_data/delivery \ $SBEAMS/var/Microarray/Affy_data/annotation \ $SBEAMS/tmp/Microarray/Make_MEV_jws_files \ $SBEAMS/tmp/Microarray/Make_MEV_jws_files/jws chgrp -R sbeams $SBEAMS/tmp/Microarray; chgrp -R sbeams $SBEAMS/var/Microarray; chmod -R g+ws $SBEAMS/tmp/Microarray; chmod -R g+ws $SBEAMS/var/Microarray; ------------------------------------------------------------------------------- 4) Edit the SBEAMS Configuration files cd $SBEAMS/lib/conf edit SBEAMS.conf Specifically: # In the DBPREFIX section, change to the correct db name, see main # sbeams.installnotes for more information DBPREFIX{Microarray} = microarray.dbo. # Next, the general microarray settings are grouped under 'Microarray Settings' # The section below tells a bit about each setting. Creating the directories # specified above and leaving the default settings should result in a working # system. # URL to help docs index page, if applicable CONFIG_SETTING{MA_AFFY_HELPDOCS_URL} # Is set to true value, 2-color menus and options will not be shown CONFIG_SETTING{MA_HIDE_TWO_COLOR} = 0 # Specify paths to R executable and library CONFIG_SETTING{MA_R_EXE_PATH} = /usr/local/bin/R CONFIG_SETTING{MA_R_LIB_PATH} = /usr/local/lib/R/library # The next 6 settings below can be specified as absolute or relative paths, see # RELATIVE VS. ABSOLUTE PATHS below for details. Default settings are relative # Directory to which affy log files will be written, if not set will default # to LOG_BASE_DIR above, else $sbeams/var/log. CONFIG_SETTING{MA_LOG_BASE_DIR} = var/logs # Full path to a folder that will house all Affy CEL files and support files CONFIG_SETTING{MA_AFFY_PROBE_DIR} = var/Microarray/Affy_data/probe_data # Location that all analysis results will be written to(see below) CONFIG_SETTING{MA_BIOC_DELIVERY_PATH} = var/Microarray/Affy_data/delivery # Location that errors from loading affy annotation files will be kept CONFIG_SETTING{MA_ANNOTATION_OUT_PATH} = tmp/Microarray/Affy_data/annotation # path to tmp dir CONFIG_SETTING{MA_AFFY_TMP_DIR} = tmp/Microarray # Location where zip files will be created for user downloads CONFIG_SETTING{MA_AFFY_ZIP_REQUEST_DIR} = tmp/Microarray/zip_request_dir # files that will be used to determine if an entire group of files, all sharing # the same basename, are present when uploading Affy arrays CONFIG_SETTING{MA_AFFY_DEFAULT_FILES} = CHP CEL XML INFO RPT R_CHP JPEG EGRAM_PF.jpg EGRAM_T.jpg EGRAM_F.jpg # Current protocol that describes the R script to produce the CHP like file $CONFIG_SETTING{MA_AFFY_R_CHP_PROTOCOL} = R Mas5.0 CHP # Notes: The web server user will need read access to the CEL file folders and # read/write access to the $CONFIG_SETTING{MA_BIOC_DELIVERY_PATH} folder ------------------------------------------------------------------------------- 5) Populate the driver tables and register the module cd $SBEAMS/lib/scripts/Core set CONFDIR = "../../conf" ./update_driver_tables.pl $CONFDIR/Microarray/Microarray_table_property.txt ./update_driver_tables.pl $CONFDIR/Microarray/Microarray_table_column.txt ./update_driver_tables.pl $CONFDIR/Microarray/Microarray_table_column_manual.txt If this doesn't work. Do not proceed, debug first. -Register the module: $SBEAMS/lib/scripts/Core/addModule.pl Microarray ------------------------------------------------------------------------------- 6) Add required Microarray work_groups and TGS entries. $SBEAMS/lib/scripts/Core/DataImport.pl -s $SBEAMS/lib/refdata/Microarray/Microarray_work_groups.xml You should now see a link for Microarray in the webUI (may require that you log back in). In order to use some of the administrative features you will have to add yourself to one or more of the work_groups that you created in the command above, Microarray_admin and Microarray_user. With your browser, navigate to: [SBEAMS Home] [Admin] [Manage User Group Associations], [Add ...], and add yourself (and appropriate others) to these groups. Now that the Microarray driver tables are loaded, and the groups have been established, you should be able to go to the web site again and click on SBEAMS - Microarray and explore the tables. They're all going to be empty, but you shouldn't get any errors, just empty resultsets. If this doesn't work. Do not proceed, debug first. ------------------------------------------------------------------------------- 7) Add some sample data ######################## Test Data Background - Set 1) Set of 12 HG-U133A arrays to demonstrate loading externally-generated data: External_test_data.tar.gz - Set 2) 4 arrays on (HG-133 Plus 2) to be loaded as internally-generated data. Experiments with C4-2 and LNCaP human prostate cancer cell lines. File: Affy_test_data.tar.gz ######################## - Download the data sets from http://www.sbeams.org/sample_data/ Click on the link "External_test_data.tar.gz" Click on the link "Affy_test_data.tar.gz" Save the data to any folder or use the folder $SBEAMS/lib/refdata/Microarray Alternatively, cd to the target directory and fetch the files with wget: wget http://www.sbeams.org/sample_data/Microarray/External_test_data.tar.gz wget http://www.sbeams.org/sample_data/Microarray/Affy_test_data.tar.gz You can check the integrity of the downloads with the md5sum command: >md5sum External_test_data.tar.gz b77678175f9bbe6aba67398a1c34fcb1 External_test_data.tar.gz >md5sum Affy_test_data.tar.gz 04d81273f454701e74526b450e79b740 Affy_test_data.tar.gz Check the download website for the most up-to-date checksum values. - Unpack the data and annotation file and copy them to the data folders cd $SBEAMS/lib/refdata/Microarray tar xvfz External_test_data.tar.gz tar xvfz Affy_test_data.tar.gz ## Note that the directories below are based on the default directory structure, ## you may need to modify the paths if you've changed anything. -Copy the annotation file to annotation directory cp External_test_data/HG-U133A_annot.csv $SBEAMS/var/Microarray/Affy_data/annotation/ cp Affy_test_data/HG-U133_Plus_2_annot.csv $SBEAMS/var/Microarray/Affy_data/annotation/ - Make the folders to hold the CEL files mkdir $SBEAMS/var/Microarray/Affy_data/probe_data/external/External_test mkdir $SBEAMS/var/Microarray/Affy_data/probe_data/200404 - Copy the the CEL files and info file to the test folder cp External_test_data/* $SBEAMS/var/Microarray/Affy_data/probe_data/external/External_test/ -Copy All the files CEL, XML, RPT files from the Affy_test_data to the data folder cp Affy_test_data/* $SBEAMS/var/Microarray/Affy_data/probe_data/200404/ ######################## (logged in as your user account) Add a contact as follows: - Switch to the Admin group by using the drop-down box at top - Click on [SBEAMS Home] - Click on [Admin] - Click [Manage Contacts] and [Add Contact] - Make a new contact with the following information: Last Name: Stegmaier First Name: Kimberly Contact Type: data_producer Organization: UNKNOWN - Fill in the appropriate information and click [INSERT] Add a user login as follows: - Switch to the Admin group by using the drop-down box at top - Click on [SBEAMS Home] - Click on [Admin] - Click on [Add User Login] - Make a new user login with the following information: Contact: Stegmaier, Kimberly Username: kstegmaier Privelege: data_reader - Fill in the appropriate information and click [INSERT] Add a Project: - Switch to the Microarray_user group by using the drop-down box at top - Click on [SBEAMS Home] - Under "My Projects" tab, click [Add A New Project]. Required fields are in red. If you don't have a budget number, enter NA. - Make a project using the following information Project Name: "Primary APL Samples" Project Tag: "PrimaryAPL" PI Contact: "Stegmaier, Kimberly" Description: "Human cell type arrays from Stegmaier 2004 paper." Budget: "NA" - Fill in the appropriate information and click [INSERT] Add a second Project: - Click the 'Go Back' button - Make a project using the following information Project Name: "ISB Test Data" Project Tag: "ISB_test" Description: "4 Human CEL files HG-U133_Plus_2. 2 files from Human LNCaP prostate cancer cell, 2files from C4-2 cancer cell lines" - Fill in the appropriate information and click [INSERT] ######################## Add An Array Type - Switch to the Microarray_admin group by using the drop-down box at top - Switch to the the Microarray Module, Click On [Microarray] - At the bottom of the page, on the Navigation bar click on [Slide Types] button - Click On [Add Slide Type] - Fill in the Data Name: "HG-U133A" Organism: "Human" Comment: "Affymetrix Human Genome U133A Array" - Click Insert - Make a second array - Click the 'Go Back' Button Name: "HG-U133_Plus_2" Organism: "Human" Comment: "Affymetrix Human Genome U133 Plus 2.0 Array" - Click Insert ######################## Add Analysis Protocol - Switch to the Microarray_admin group by using the drop-down box at top - Switch to the the Microarray Module, Click On [Microarray] - On the Navigation bar click on [Protocols] button - Click [Add Protocol] - Add a new Protocol Type, Click the "Plus Icon" next to Protocol Type - Fill in Protocol Type Name --> "image_analysis" - Click Insert. - Click "Go Back" Button - Insert the additional Protocol types "hybridization" "array_scanning" - Go Back to the Add Protocol Page, and refresh the page. - Insert the data Protocol Type --> "image_analysis", Name --> "R Mas5.0 CHP", Protocol --> "Use R-bioconductor to turn a CEL file into a CHP file" - Click Insert ######################## Add Hyb Protocol - Click "Go Back" Button - Insert the data Protocol Type --> "hybridization", Name --> "AFFY EukGE-WS2v5_450", Protocol --> "Paste in the text output from the affy wash station" - Click Insert ######################## Add Scanning Protocol - Click the "Go Back" Button - Insert the data Protocol Type --> "array_scanning", Name --> "AFFY Scanning", Protocol --> "Paste in the text output from the affy scanner" - Click Insert ######################## Add Feature extraction Protocol - Click the "Go Back" Button - Insert the data Protocol Type --> "image_analysis", Name --> "AFFY Feature Extraction", Protocol --> "Generation of CEL file from DAT file is automatic in GCOS 1.0+ Software" - Click Insert ######################## Add CHP Generation Protocol - Click the "Go Back" Button - Insert the data Protocol Type --> "image_analysis", Name --> "AFFY GCOS CHP Generation", Protocol --> "Create CHP file in GCOS by right-clicking a CEL file and choosing 'Analyze'" - Click Insert ######################## Add Server Info - Some of the scripts and database tables keep track of file paths and the server name needs to be entered before some of the scripts will work - Manually construct the URL to enter the data below - The url should be almost the same for the ones above for adding an array, but the TABLE_NAME will = MA_server http:///sbeams/cgi/Microarray/ManageTable.cgi?TABLE_NAME=MA_server - Enter the server name - Click [Insert] ######################## Add the Annotation for this array type - Annotations are downloaded from Affymetrix on quarterly basis, and must be done manually. The file provided in the test data set was downloaded from Affymetrix. - Use the script to upload the annotation into SBEAMS. These are large files, - and the process could take 20 minutes or more per file. Note that the - Plus 2 array has about twice as many probsets as the 133A. cd $SBEAMS/lib/scripts/Microarray ./load_affy_annotation_files.pl --run_mode update \ --file_name $SBEAMS/var/Microarray/Affy_data/annotation/HG-U133A_annot.csv - Add the annotation for the second array ./load_affy_annotation_files.pl --run_mode update \ --file_name $SBEAMS/var/Microarray/Affy_data/annotation/HG-U133_Plus_2_annot.csv ######################## Add the external arrays to the database - See sections about naming conventions (3-2) - The Data in the "External_test_data.tar.gz" folder contains CEL files that were not generated at ISB, so they did not have all the support data needed to automatically load the arrays from the core facility. - Adding external arrays is a two step process. - All the sample annotation is first collected in a "master_info" file which is just a tab delimited file. - This file is then parsed to produce a info_file for each CEL file to be uploaded - See Below on how to upload data from a MAGE-XML file produced by the Affymetrix GCOS software - View the template on how to make a master_info file. [Not needed for this example, the info file already exists] $SBEAMS/usr/Microarray/Example_master_upload_template.xls - Run the script to parse the master_info file cd $SBEAMS/lib/scripts/Microarray ./Pre_process_affy_info_file.pl --run_mode make_new \ --info_file $SBEAMS/var/Microarray/Affy_data/probe_data/external/External_test/External_test_data_master.txt - Run the script to upload the data into SBEAMS - This script will scan the default data directory (set in the Settings file) looking for new data cd $SBEAMS/lib/scripts/Microarray; ./load_affy_array_files.pl --run_mode add_new ######################## Add more data to the database MAGE-XML data. - The data contained within the file "Affy_test_data.tar.gz" contains from 4 arrays scanned at ISB. - In addition to the CEL file is a CHP, RPT and XML files that were produced by the Affymetrix software GCOS. - ISB exports the MAGE-XML from the GCOS software with one extra attribute that will be needed for the current script to parse the data. - Setup GCOS software ## Only do this if you have an affy core facility and you want to load the data they produced Goto the GCOS Manager, Click on the template tab. Create a new template Add New Attribute "Array User Name" Type "String" Required "Yes" Make sure to use the template when exporting data from the GCOS software after scanning slides ######################## Add expression data to the database Convert CEL file to CHP data using R-bioconductor cd $SBEAMS/lib/scripts/Microarray; ./load_affy_R_CHP_files.pl --run_mode add_new ##Note this will take a while of time to process, and running this on a fast machine would be a good idea - Test to see if the data has been entered properly - Open the url http:///sbeams/cgi/Microarray/GetAffy_GeneIntensity.cgi - Enter "IL%" in the box Gene Name, Click 'Simple Query' ------------------------------------------------------------------------------- ######################## 8) Setup the help pages ######################## Setting up the Affy user Help pages - There is a set of user help pages that were made for the end users and it shows how to use the different pages - The pages need an XSL processing engine to work. Currently they are setup to use xsltproc by default. Check to to see if xsltproc is installed xsltproc -V If it's not installed goto http://xmlsoft.org/XSLT/ and install the programs - Set the site specific information to locate the files. Go and edit the following two files. cd $SBEAMS/doc/Microarray/affy_help_pages/includes - Edit the file vi transformation_info.inc.php - Change the following variables to point to where the files are installed Example, Change $SBEAMS to suit your needs $XSLT_URL = "http://<$SBEAMS>/doc/Microarray/Affy_help/Affy_help.xslt"; $XML_URL = "http://<$SBEAMS>/doc/Microarray/Affy_help"; $URL_BASE = "http://<$SBEAMS>/doc/Microarray/affy_help_pages" $BASE_PATH = "<$SBEAMS>/doc/Microarray/affy_help_pages/Affy_help" $HOME_PAGE_URL = "" - Edit the xslt file to add the correct server name cd $SBEAMS/doc/Microarray/affy_help_pages/Affy_help vi Affy_help.xslt - Change the lines, to indicate your servers that are running both SBEAMS and the affy help pages db.systemsbiology.net db.systemsbiology.net To See if the links work check out the page: http:///doc/Microarray/affy_help_pages/ Most of the links are relative but a few links go directly into SBEAMS therefore the need to set the above links. Also note some of the links will not work since they are specific to ISB ------------------------------------------------------------------------------- ############################## 9) Setup the analysis pipeline ############################## The analysis pipeline allows users to analyze affy arrays and find differentially expressed genes. The analysis is done in R and the processing can be done on the local server or passed off to a batch processor like pbs. A series of web pages will walk the users through the analysis pipeline so there is no need for the user to read/write R code. The CGI pages were initially written as part of the bioconductor project. See the bioconductor vignette on "Textual Description of webbioc" to learn more about the requirements for running and requirements for running these pages (Link below). It should be noted that you do NOT have to download the pages from bioconductor, the pages have been total integrated into SBEAMS so the actual use of the pages is a bit different and the analysis capabilities have been extended, but the setup requirements and some of the settings are still applicable http://bioconductor.org/repository/devel/vignette/demoscript.pdf # Create a new unix group, affydata, for example: /usr/sbin/groupadd affydata # Add web user to this group, e.g. (assumes web server runs as 'apache'): /usr/sbin/usermod -G affydata apache # Change the group of the delivery dir specified above (MA_BIOC_DELIVERY_PATH), # default locatin used here as an example. chgrp -R affydata $SBEAMS/var/Microarray/Affy_data/delivery # Set sticky bit so all data will have the group arraydata chmod g+s $SBEAMS/var/Microarray/Affy_data/delivery You can run the bioconductor jobs as a forked process, or farm them to a batch scheduler such as pbs. To run the job on pbs you may need to create a new user (See Below), and set up a keyless ssh connection apache cgi script can hand the job off to the batch scheduler, and that process can write files back. This is beyond the scope of this document, but we can provide more info if desired. In a default bioconductor installation, site specific settings are specified in the Site.pm module, located within SBEAMS at; $SBEAMS/cgi/Microarray/bioconductor/Site.pm Virtually all such settings can now be specified in the SBEAMS.conf file, as outlined below. One exception is the 'batch type' as described above. If an option other than fork is chosen (the default in SBEAMS.conf), you will need to edit Site.pm directly. # Affy admin email address. Notification of jobs being completed is sent to # users if they request it; this is the 'senders address'. CONFIG_SETTING{MA_ADMIN_EMAIL} = sbeams@localdomain.org # Specify paths to R executable and library, done previously. # Batch method for running bioconductor R jobs. Valid options include fork, # pbs, and sge. pbs and sge will require some source file editing to set up. CONFIG_SETTING{MA_BATCH_SYSTEM} = fork # path to bioconductor delivery directory. This is where various script and # output files are stored, default given as a path relative to sbeams root. CONFIG_SETTING{MA_BIOC_DELIVERY_PATH} = var/Microarray/Affy_data/delivery # path to affy annotation files. These are tab-delimited text files from # affymetrix describing the gene that each probeset represents. CONFIG_SETTING{MA_ANNOTATION_PATH} = var/Microarray/Affy_data/annotation # Settings for java/jnlp helper applications. These are required to use the # packaged cytoscape or Tigr MEV (multi-array-viewer) applications. Leaving # the defaults should result in a working system; if you would like to change # the certificate to a local version, you will have to unjar and rejar each of # the files under $sbeams/lib/java/SBEAMS/Microarray (see OPTIONAL section # below) CONFIG_SETTING{JAVA_PATH} = /usr/java/j2sdk1.4/ CONFIG_SETTING{JNLP_KEYSTORE} = lib/java/.keystore CONFIG_SETTING{KEYSTORE_PASSWD} = sbeams_distro CONFIG_SETTING{KEYSTORE_ALIAS} = sbeamsDistro ### OPTIONAL - create site-specific keystore and re-sign jar files #### # Setup the a keystore for sbeams, for example $JAVA_PATH/bin/keytool -keystore $JNLP_KEYSTORE -genkey -alias KEYSTORE_ALIAS Enter keystore password: sbeamsDevKey What is your first and last name? [Unknown]: SBEAMS Development Team What is the name of your organizational unit? [Unknown]: SBEAMS Development What is the name of your organization? [Unknown]: Institute for Systems Biology What is the name of your City or Locality? [Unknown]: Seattle What is the name of your State or Province? [Unknown]: WA What is the two-letter country code for this unit? [Unknown]: US Is CN=SBEAMS Development Team, OU=SBEAMS Development, O=Institute for Systems Biology, L=Seattle, ST=WA, C=US correct? [no]: yes Enter key password for <$KEYSTORE_ALIAS> (RETURN if same as keystore password): # Re-sign the all the cytoscape jars using the key above cd $SBEAMS/lib/scripts/Core # Re-sign all the cytoscape and gaggle components ./Re_sign_jar_files.pl --dir $SBEAMS/usr/java/share/Cytoscape/ ./Re_sign_jar_files.pl --dir $SBEAMS/usr/java/share/Cytoscape/cytoscape_ps/jars # Re-sign all the plugin jars ./Re_sign_jar_files.pl --dir $SBEAMS/usr/java/share/Cytoscape/plugins_1.0 ./Re_sign_jar_files.pl --dir $SBEAMS/usr/java/share/Cytoscape/plugins_2.0 ### END OPTIONAL keystore setup section ### To run the pages #Open the main Microarray Module web page - Click on [Microarray] - Click on [Data Pipeline] - Click on [Start New Analysis Session] #Make sure there are no errors displayed, if there are go back and check the settings. View the help document to learn how to analyze data sets http:///doc/Microarray/affy_help_pages/isb_help.php?help_page=Analysis/Pipeline/Pipeline_overview.xml Read about, "Grouping arrays","Normalizing Array Data" and "Analyze Arrays" ------------------------------------------------------------------------------- Troubleshooting Problem: Analysis Pipeline doesn't produce diagnostic PNG images during normalization, and the R error code file shows "Ghostscript *.**: Unrecoverable error, exit code 1". Alternatively, producing JPEG images from the chips during load_affy_R_CHP_files.pl runs may not work. Fix: Install AFPL Ghostscript 8.51. Problem: Analysis Pipeline produces truncated (size 0) png files during normalization and SAM/ttest analysis. Fix: Netpbm libraries not installed or not in path of analysis user (arraybot), download and install from http://netpbm.sourceforge.net