Annotation pipeline
===================

This Annotation pipeline is an automatic structural annotation workflows written in snakemake. The annotation is based on two tools which uses RNA-Seq and/or protein homology information for predict coding sequence. One of this tools is BRAKER [ref & lien] which use GeneMark-EX [ref] and AUGUSTUS [ref]. And the other tool is Augustus [ref] alone for improve annotation of small coding sequences with few or no intron. 


.. image:: image/schema_pipeline.jpg
   :target: image/schema_pipeline.jpg
   :width: 300
   :align: center
   :alt: global


Mandatory installation
----------------------


- \ `Hisat2 <https://ccb.jhu.edu/software/hisat2/manual.shtml#obtaining-hisat2>`_\ 
- \ `samtools <https://github.com/samtools/samtools>`_\ 
- \ `exonerate <https://www.ebi.ac.uk/about/vertebrate-genomics/software/exonerate>`_\ 
- \ `BRAKER <https://github.com/Gaius-Augustus/BRAKER>`_\ 
- \ `BUSCO <https://gitlab.com/ezlab/busco/-/tree/master>`_\ 
- \ `R <https://cran.r-project.org/bin/linux/ubuntu/README.html>`_ & `Rmarckdown package <https://rmarkdown.rstudio.com/lesson-1.html>`_\ **
- \ `Python >=3.7 <https://www.python.org/downloads/>`_\ 
- \ `Snakemake >= 5.2 <https://snakemake.readthedocs.io/en/stable/>`_\ 

Prepare config file
-------------------

To run the workflows you have to provide the data path for all input file. Please complete the config.yaml file for launch the workflow. 

1. Providing data
^^^^^^^^^^^^^^^^^

.. code-block::

   {
           "datadir"   : "/homedir/user/work/Masked_assembly/",
           "OutDir"    : "/homedir/user/work/pipeline_output/",
           "RnaSeqDir" : "/homedir/user/work//rnaseq/",
           "id_souche" : "magnaporthe_oryzae",
           "protRef"   : "/homedir/user/work/Annotation_pipeline/SupplementaryFile/OG_filter.fasta"
   }


- **datadir** : Path of you directory which contain all your fasta file to annotate. 
- **RnaSeqDir** : Path of the directory which contain all RNAseq data,
- **id_souche** : ID of species for augustus trainings, please refers to augustus main page for this option
- **protRef** :  Path of the protein fasta file.
- **OutDir** : Output directory for all results of this pipeline

Launching on a single machine
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^

To launch this annotation pipeline on a single machine, please use the command :

.. code-block::

   snakemake -s annotation_pipeline.snake --latency-wait 555555

2. Launching on HPC clusters
^^^^^^^^^^^^^^^^^^^^^^^^^^^^

Preparing cluster configuration using cluster_config.yaml
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

On cluster_config.yaml, you can add partition, memory and to be used by default for each rule. This pipeline is used on cirad HPC [link], so the environment (module load command) and sge submission works only on the cirad cluster. Please adapt the content of this file before running on a cluster with your cluster parameters or if you need more memory for a rules. 

Here is a example of the configuration file we used on the Cirad HPC.

.. code-block::

   __default__:
     n_cpu: "-pe parallel_smp"
     mem: "-l mem_free=4G"
     export_env: "-V"
     cwd: "-cwd"
     logerror: "-e "
     log: "-o "
     queue: " -q long.q"
     scheduler: "qsub"

   ###### Cluster parameters for hisat2_index rules ########
   hisat2_index :
     mem: "-l mem_free=4G"
     queue: " -q normal.q"

Launch workflows
~~~~~~~~~~~~~~~~

For launch the pipeline in a cluster please use the **Launcher.sh** give in the directory.