<tool id="ribotaper_ribosome_profiling" name="ribotaper part 3: ribosome profiling" version="0.1.0">
    <requirements>
            <requirement type="package" version="1.3.1">ribotaper</requirement>
    </requirements>
    <stdio>
        <exit_code range="1:" />
    </stdio>

    <command><![CDATA[
        tar
            "xzvf"
            "$annotation_path"

        &&

        Ribotaper.sh
            "$ribo_bam"
            "$rna_bam"
            "annotation_path"
            "$read_lenghts_ribo1,$read_lenghts_ribo2,$read_lenghts_ribo3"
            "$cutoff1,$cutoff2,$cutoff3"
            "\${GALAXY_SLOTS:-12}"

    ]]></command>
    <inputs>
        <param name="annotation_path" type="data" format="compressed_archive" label="annotation_path" help="Please run 'ribotaper part 1' to generate the archive."/>
        <param name="ribo_bam" type="data" format="BAM" label="ribo_bam" help="Ribo-seq alignment file in BAM format."/>
        <param name="rna_bam" type="data" format="BAM" label="rna_bam" help="RNA-seq alignment file in BAM format."/>
        <param name="read_lenghts_ribo1" type="text" value="26"  label="Read length 1" help="Read length 1, which is used for P-site calculation. Default is '26' but it varies a lot in different datasets. Please run 'ribotaper part 2' to deterimine a appropriate value."/>
        <param name="read_lenghts_ribo2" type="text" value="28"  label="Read length 2" help="Read length 2, which is used for P-site calculation. Default is '28' but it varies a lot in different datasets.  Please run 'ribotaper part 2' to deterimine a appropriate value"/>
        <param name="read_lenghts_ribo3" type="text" value="29"  label="Read length 3" help="Read length 3, which is used for P-site calculation. Default is '29' but it varies a lot in different datasets.  Please run 'ribotaper part 2' to deterimine a appropriate value"/>
        <param name="cutoff1" type="text" value="9"  label="Cutoff 1" help="Offset 1, which is used for P-sites calculation. Default is '9' but it varies a lot in different datasets.
        Please run 'ribotaper part 2' to deterimine a appropriate value."/>
        <param name="cutoff2" type="text" value="12"  label="Cutoff 2" help="Offset 2, which is used for P-sites calculation. Default is '12' but it varies a lot in different datasets.
         Please run 'ribotaper part 2' to deterimine a appropriate value."/>
        <param name="cutoff3" type="text" value="12"  label="Cutoff 3" help="Offset 3, which is used for P-sites calculation. Default is '12' but it varies a lot in different datasets.
         Please run 'ribotaper part 2' to deterimine a appropriate value."/>
    </inputs>
    <outputs>
        <data name="output1" type="data" format="pdf" from_work_dir="quality_check_plots.pdf" label="QC plots"/>
        <data name="output2" type="data" format="tabular" from_work_dir="ORFs_genes_found" label="Summary of translated ORFs"/>
        <data name="output3" type="data" format="tabular" from_work_dir="ORFs_max" label="Translated ORFs (max)"/>
        <data name="output4" type="data" format="tabular" from_work_dir="ORFs_max_filt" label="Translated ORFs (max_filt)"/>
        <data name="output5" type="data" format="bed" from_work_dir="translated_ORFs_sorted.bed" label="Translated ORFs (sorted)"/>
        <data name="output6" type="data" format="bed" from_work_dir="translated_ORFs_filtered_sorted.bed" label="Translated ORFs (filtered/sorted)"/>
        <data name="output7" type="data" format="fasta" from_work_dir="protein_db_max.fasta" label="Protein DB"/>
        <data name="output8" type="data" format="pdf" from_work_dir="Final_ORF_results.pdf" label="ORF categories (length/coverage)"/>
    </outputs>
    <tests>
        <test>
            <param name="annotation_path" value="annotation_path.tgz" ftype="compressed_archive"/>
            <param name="ribo_bam" value="test_ribo.bam"/>
            <param name="rna_bam" value="test_rna.bam"/>
            <param name="read_lenghts_ribo1" value="26"/>
            <param name="read_lenghts_ribo2" value="28"/>
            <param name="read_lenghts_ribo3" value="29"/>
            <param name="cutoff1" value="9"/>
            <param name="cutoff2" value="12"/>
            <param name="cutoff3" value="12"/>
            <output name="output2" file="ORFs_genes_found"/>
        </test>
    </tests>
    <help><![CDATA[
RiboTaper is an analysis pipeline for Ribosome Profiling
(Ribo-seq) experiments,
which exploits the triplet periodicity of
ribosomal footprints to call translated regions.
See
https://ohlerlab.mdc-berlin.de/software/RiboTaper_126/ for details.


The Ribotaper Galaxy tool set consists of three tools:

  - ``ribotaper part 1``: creation of annotation files
  - ``ribotaper part 2``: metagene analysis for P-sites definition
  - ``ribotaper part 3``: ribosome profiling

The order of execution should follow:
``ribotaper part 1, part 2 and part 3``.

The current tool is ``ribotaper part 3``,
ribosome profiling.

Outputs
--------

**QC plots**:
This plot provides the user statistics about the Ribo-seq and RNA seq data used, together with the assessment of the P-sites calculations.
Important values are the pie chart showing the agreement between the frame (defined by the P-sites position) and the annotated frame. Reliable P-sites calculations produce an agreement above 90%.
Very important are also the length/coverage statistics for the Ribo-seq (bottom right):
This shows how the P-site calculations can be used to detect active translation in regions of different length and coverage, in a way the user can estimate the precision of the Ribo-seq data, and understand the level or resolution the data allows.

**Summary of translated ORFs**:
Tab-separated values for the number of ORFs found and their corresponding genes, for the different ORF categories.

**Translated ORFs (max, max_filt)**:
Tab-separated file containing information about detected ORFs.
Translated ORFs (max_filt) contains ORFs filtered for excessive multimapping and ORFs in non-coding genes overlapping known coding regions (recommended for further analysis).

**Protein DB**:
Fasta file of the detected ORFs peptide sequence, suitable as an alternative protein database (not filtered for multimapping)

**Translated ORFs  (sorted, filtered/sorted)**:
BED files with genomic coordinates for the detected ORFs. The total number of P-sites along the ORF is reported on the 5th column.

**ORF categories (length/coverage)**:
PDF file containing info about the number of ORFs found, together with their length and coverage per category/annotation.

Important notes
----------------

  - We ran the RiboTaper analysis on an SGE cluster, using 7 cores and h_vmem 8G. For each dataset, the complete RiboTaper workflow (from the bam files to final results) took ~ 1 day.

  - The current RiboTaper framework is not designed to identify and quantify ORFs on different transcripts. This means the transcript annotation is crucial.

  - Be careful about using scaffolds, both in the genome and GTF files, which may slow the whole pipeline.

]]></help>
    <citations>
        <citation type="doi">10.1038/nmeth.3688</citation>
    </citations>
</tool>