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Gene Set Clustering based on Functional annotation. Most up-to-date and realtime information based gene enrichment analysis.
Publication:
GeneSCF: a real-time based functional enrichment tool with support for multiple organisms. BMC Bioinformatics 17, 365 (2016). https://doi.org/10.1186/s12859-016-1250-z

This documentation will provide detailed information on usage of GeneSCF tool (All versions). Read following page for running GeneSCF on test dataset.

• Overview
• Installation
• General usage
• Step-by-step instructions (simple usage)
  • Preparing database
  • Enrichment analysis
  • Single-step enrichment analysis
• GeneSCF batch analysis
  • Preparing arguments file
  • Edit batch script
  • Run batch analysis
• License

GeneSCF is a command line based and powerful tool to perform gene enrichment analysis. GeneSCF uses realtime informatin from repositories such as geneontology, KEGG, Reactome and NCG while performing the analysis. This increases reliability of the outcome compared to other available tools. GeneSCF is command line tool designed to easily integrate with any next-generation sequencing analysis pipelines. One can use multiple gene list in parallel to save time. In simple terms,

• Real-time analysis, do not have to depend on enrichment tools to get updated.
• Easy for computational biologists to integrate this simple tool with their NGS pipeline.
• GeneSCF supports more organisms.
• Enrichment analysis for Multiple gene list in single run.
• Enrichment analysis for Multiple gene list using Multiple source database (GO,KEGG, REACTOME and NCG) in single run.
• Download complete GO terms/Pathways/Functions with associated genes as simple table format in a plain text file. Check Preparing database under Step-by-step instructions (simple usage) section for the details.

Above graphs are from Subhash and Kanduri et al., 2016

Download and extract the compressed file using 'unzip' (for .zip file) or 'tar' (for .tar.gz file). Use it without any need for special installation procedures.

• Dependencies: PERL
  
• Test required modules : awk, cat, unzip, gzip, wget, rm, mkdir, sort, date, sed, paste, join, grep, curl, echo
  
• For graphical output or plots: If needed graphical output, pre installated R (version > 3.0) and 'ggplot2' is required.
  Note: GeneSCF only works on Linux system, it has been successfully tested on Ubuntu, Mint and Cent OS. Other distributions of Linux might work as well.

There are two utilities available from GeneSCF package.

1. One is the main command line 'geneSCF', to perform gene enrichment analysis.
2. Next, is the 'prepare_database' command line to prepare the necessary database of an organism. GeneSCF by default comes with database for human consists of gene ontology, KEGG, Reactome, and NCG.

Note: Replace 'geneSCF-master-vx.x' from the commands with your GeneSCF directory.

./geneSCF-master-vx.x/geneSCF -m=[update|normal] -i=[INPUT FILE] -t=[gid|sym] -o=[OUTPUT PATH/FOLDER/] -db=[GO_all|GO_BP|GO_MF|GO_CC|KEGG|REACTOME] -p=[yes|no] -bg=[#TotalGenes] -org=[see,org_codes_help]

./geneSCF-master-vx.x/prepare_database -db=[GO_all|GO_BP|GO_MF|GO_CC|KEGG|REACTOME] -org=[see,org_codes_help directory]

Note: The above command downloads complete '-db' of your choice as simple text file with corresponding genes per GO term in following location, 'geneSCF-master-vx.x/class/lib/db/[ORGANISM]/' for your prefered organism.

For a convenience we will use test datasets from the directory 'geneSCF-master-vx.x/test/'. There are two steps involved,

1. Prepare your prefered database for an organism of your interest.
2. Perform enrichment analysis on your gene list.
3. One can also perform enrichment analysis in single-step using 'update' mode.

./geneSCF-master-vx.x/prepare_database -db=GO_all -org=goa_human

Note: Specific dabases can be also updated using 'GO_BP', 'GO_MF' and 'GO_CC'. The above command downloads complete geneontology ('GO_all') with corresponding genes per GO term as simple text file in following location, 'geneSCF-master-vx.x/class/lib/db/goa_human/'.

./geneSCF-master-vx.x/prepare_database -db=KEGG -org=hsa

./geneSCF-master-vx.x/prepare_database -db=REACTOME -org=Hs

Note: Reactome supports only Human (Hs)

./geneSCF-master-vx.x/prepare_database -db=NCG -org=Hs

Note: NCG supports only Human (Hs)

./geneSCF-master-vx.x/geneSCF -m=normal -i=geneSCF-master-vx.x/test/H0.list -o=geneSCF-master-vx.x/test/output/ -t=sym -db=GO_BP -bg=20000 --plot=yes -org=goa_human

./geneSCF-master-vx.x/geneSCF -m=normal -i=geneSCF-master-vx.x/test/H0.list -o=geneSCF-master-vx.x/test/output/ -t=sym -db=KEGG -bg=20000 --plot=yes -org=hsa

Note: All predicted results can be found in 'geneSCF-master-vx.x/test/output/' folder with file name '{INPUT_FILE_NAME}_{database}_functional_classification.tsv'

This '-m=update' mode will integrate both 'prepare_database' and 'geneSCF' into single command mode. When you use 'update' mode once, you can use 'normal' mode for the next consecutive runs, in case you are planning to use the same database for different gene lists (This saves time).

./geneSCF-master-vx.x/geneSCF -m=update -i=geneSCF-master-vx.x/test/H0.list -o=geneSCF-master-vx.x/test/output/ -t=sym -db=GO_BP -bg=20000 --plot=yes -org=goa_human

**Note:** The above command also downloads complete geneontology biological processes ('GO_BP') with corresponding genes per GO term as a simple text file in following location, '<b>geneSCF-master-vx.x/class/lib/db/goa_human/</b>' and also does enrichment analysis in parallel. The results for enrichment analysis can be found in folder '<b>geneSCF-master-vx.x/test/output/</b>'.

./geneSCF-master-vx.x/geneSCF -m=update -i=geneSCF-master-vx.x/test/H0.list -o=geneSCF-master-vx.x/test/output/ -t=sym -db=KEGG -bg=20000 --plot=yes -org=hsa

Note: All predicted results can be found in 'geneSCF-master-vx.x/test/output/' folder with file name '{INPUT_FILE_NAME}_{database}_functional_classification.tsv'

Edit file './geneSCF-master-vx.x/db_batch_config.txt' to configure your parameters for batch run. The sample file looks like the one below,

#database:organism:background:type
#GO_BP:goa_human:20000:sym
#GO_MF:goa_human:20000:sym
GO_CC:goa_human:20000:sym
KEGG:hsa:20000:sym
#REACTOME:Hs:20000:sym
NCG:Hs:20000:sym

In the above file you are asking GeneSCF to run enrichment analysis using 'GO_CC', 'KEGG', and 'NCG' database for human. The database mentioned with preceeding '#' will not be considered for the run.

Edit script './geneSCF-master-vx.x/geneSCF_batch' for your input files (files_path) and output path (output_path).

files_path="/FOLDER/WHERE/GENE_LISTS/STORED"
output_path="/FOLDER/PATH/FOR/OUTPUT"

Note:

• It is recommended to keep all input files in same folder.
• Inside specified output folder path GeneSCF will automatically create individual sub-folders for each gene list.

Execute batch analysis.

./geneSCF-master-vx.x/geneSCF_batch

GNU GENERAL PUBLIC LICENSE