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# RSEM Galaxy Wrapper #

## Introduction ##

RSEM (RNA-Seq by Expectation-Maximization) is a software package for the
estimation of gene and isoform abundances from RNA-Seq data. A key feature of
RSEM is its statistically-principled approach to the handling of RNA-Seq
reads that map to multiple genes and/or isoforms. In addition, RSEM is
well-suited to performing quantification with de novo transcriptome
assemblies, as it does not require a reference genome.

## Installation ##

Follow the [Galaxy Tool Shed
instructions](http://wiki.g2.bx.psu.edu/Tool_Shed) to add this wrapper from
the tool shed to your galaxy instance. Once the files are in the tools
directory you have to have RSEM references installed. This can be done by:

1. Placing the file called `rsem_indices.loc` into the directory
   `~/galaxy-dist/tool-data` This file tells the RSEM wrapper how to find the
   reference(s). It is formatted according to galaxy's documentation with the
   following tab-delimited format:

        unique_build_id    dbkey    display_name    file_base_path
	
   For example,

        human_refseq_NM	human_refseq_NM	human_refseq_NM	/opt/galaxy/references/human/1.1.2/NM_refseq_ref

2. Downloaded a pre-built RSEM reference from the [RSEM website](http://deweylab.biostat.wisc.edu/rsem/).

3. Place reference files into the `file_base_path` listed in the
`rsem_indices.loc` file

If you would rather build your own reference files follow the instructions
below and then place resulting reference files into the `file_base_path` listed
in the `rsem_indices.loc` file.

### Building a custom RSEM reference ###

For instructions on how to build the RSEM reference files, first see the [RSEM
documentation](http://deweylab.biostat.wisc.edu/rsem/README.html).

#### Example ####

Suppose we have mouse RNA-Seq data and want to use the UCSC mm9 version of the
mouse genome. We have downloaded the UCSC Genes transcript annotations in GTF
format (as mm9.gtf) using the Table Browser and the knownIsoforms.txt file for
mm9 from the UCSC Downloads. We also have all chromosome files for mm9 in the
directory `/data/mm9`. We want to put the generated reference files under
`/opt/galaxy/references` with name `mouse_125`. We'll add poly(A) tails with
length 125. Please note that GTF files generated from UCSC's Table Browser do
not contain isoform-gene relationship information. For the UCSC Genes
annotation, this information can be obtained from the knownIsoforms.txt file.
Suppose we want to build Bowtie indices and Bowtie executables are found in
`/sw/bowtie`.

To build the reference files, first run the command:

    rsem-prepare-reference --gtf mm9.gtf \
                           --transcript-to-gene-map knownIsoforms.txt \
                           --bowtie-path /sw/bowtie \                  
                           /data/mm9/chr1.fa,/data/mm9/chr2.fa,...,/data/mm9/chrM.fa \
                           /opt/galaxy/references/mouse_125

To add this reference to your galaxy installation, add the following line to
the the `rsem_indices.loc` file:

    mouse_125	mouse_125	mouse_125	/opt/galaxy/references/mouse_125

Then restart galaxy and you should see the `mouse_125` reference listed in the
RSEM wrapper.

## References ##

* [RSEM website (stand alone package)](http://deweylab.biostat.wisc.edu/rsem/)

* B. Li and C. Dewey (2011) [RSEM: accurate transcript quantification from
  RNA-Seq data with or without a reference
  genome](http://bioinformatics.oxfordjournals.org/content/26/4/493.abstract).
  BMC Bioinformatics 12:323.

* B. Li, V. Ruotti, R. Stewart, J. Thomson, and C. Dewey (2010) [RNA-Seq gene
  expression estimation with read mapping
  uncertainty](http://www.biomedcentral.com/1471-2105/12/323). Bioinformatics
  26(4): 493-500.

## Contact information ##
* RSEM galaxy wrapper questions: [e-mail]
* RSEM stand alone package questions: [e-mail]
* [RSEM announcements mailing list](http://groups.google.com/group/rsem-announce)
* [RSEM users mailing list](http://groups.google.com/group/rsem-users)