This package contains a Galaxy workflow for the detection and incorporation of single amino acid polymorphism (SAP) into a custom proteomics search database.

The workflow aligns RNA-Seq reads to the organism's genome using Tophat, call SNPs using SAMtools mpileup command, and annotates the SNPs that reside within protein-coding regions using SNPeff.
Galaxy tool "SNPeff to Peptide Fasta" generates peptide sequences with the polymorhisms.

See http://www.galaxyproject.org for information about the Galaxy Project.


Availability
============

This workflow is available to download and/or install from the main
Galaxy Tool Shed:

http://toolshed.g2.bx.psu.edu/view/galaxyp/proteomics_rnaseq_sap_db_workflow


Reference Data
==============

For Human RNAseq data this workflow was tested with a genome build named "GRCh37_canon" using reference data from:

* ftp://ftp.ensembl.org/pub/release-73/fasta/homo_sapiens/dna/Homo_sapiens.GRCh37.73.dna.chromosome.[1-9XY]*.fa.gz
* ftp://ftp.ensembl.org/pub/release-73/fasta/homo_sapiens/pep/Homo_sapiens.GRCh37.73.pep.all.fa.gz
* ftp://ftp.ensembl.org/pub/release-73/gtf/homo_sapiens/Homo_sapiens.GRCh37.73.gtf.gz


For Mouse RNAseq data this workflow was tested with a genome build named "GRCm38_canon" using reference data from:

* ftp://ftp.ensembl.org/pub/release-73/fasta/mus_musculus/dna/Mus_musculus.GRCm38.73.dna.chromosome.[1-9XY]*.fa.gz
* ftp://ftp.ensembl.org/pub/release-73/fasta/mus_musculus/pep/Mus_musculus.GRCm38.73.pep.all.fa.gz
* ftp://ftp.ensembl.org/pub/release-73/gtf/mus_musculus/Mus_musculus.GRCm38.73.gtf.gz


Genome builds "GRCh37_canon" and "GRCm38_canon" were built according to instructions in:  https://wiki.galaxyproject.org/Admin/NGS%20Local%20Setup
The builds used only the standard chromosomesi sequences from the reference fasta; the other sequences were filtered out.
The GTF was filtered to retain only those entries that referenced the standard chromosomes. 


Dependencies
============

These dependencies should be resolved automatically via the Galaxy Tool Shed:

* http://toolshed.g2.bx.psu.edu/view/devteam/tophat
* http://toolshed.g2.bx.psu.edu/view/devteam/samtools_mpileup
* http://toolshed.g2.bx.psu.edu/view/nilesh/bcftools
* http://toolshed.g2.bx.psu.edu/view/iuc/snpeff
* http://toolshed.g2.bx.psu.edu/view/jjohnson/snpeff_to_peptides

History
=======

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Version Changes
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v0.0.1  - Initial release to Tool Shed (March, 2014)
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