PAQmiR : Prediction Annotation and Quantification of miRNA with miRDeep2. The PAQmiR approach was used in the following projects:

• [1] Sunflower oil supplementation affects the expression of miR-20a-5p and miR-142-5p in the lactating bovine mammary gland. Mobuchon L, Le Guillou S, Marthey S, Laubier J, Laloë D, Bes S, Le Provost F, Leroux C. PLoS One. 2017 Dec 27;12(12):e0185511. doi: 10.1371/journal.pone.0185511

• [2] Deprivation Affects the miRNome in the Lactating Goat Mammary Gland. Mobuchon L, Marthey S, Le Guillou S, Laloë D, Le Provost F, Leroux C. PLoS One. Food 2015 Oct 16;10(10):e0140111. doi: 10.1371/journal.pone.0140111.

• [3] Annotation of the goat genome using next generation sequencing of microRNA expressed by the lactating mammary gland: comparison of three approaches. Mobuchon L, Marthey S, Boussaha M, Le Guillou S, Leroux C, Le Provost F. BMC Genomics. 2015 Apr 11;16:285. doi: 10.1186/s12864-015-1471-y.

• [4] Characterisation and comparison of lactating mouse and bovine mammary gland miRNomes. Le Guillou S, Marthey S, Laloë D, Laubier J, Mobuchon L, Leroux C, Le Provost F. PLoS One. 2014 Mar 21;9(3):e91938. doi: 10.1371/journal.pone.0091938.

• [5] A comprehensive overview of bull sperm-borne small non-coding RNAs and their diversity across breeds. Sellem E, Marthey S, Rau A, Jouneau L, Bonnet A, Perrier JP, Fritz S, Le Danvic C, Boussaha M, Kiefer H, Jammes H, Schibler L. Chromatin. 2020 Mar 30;13(1):19. doi: 10.1186/s13072-020-00340-0. PMID: 32228651; PMCID: PMC7106649.

• [6] Dynamics of cattle sperm sncRNAs during maturation, from testis to ejaculated sperm. Sellem E, Marthey S, Rau A, Jouneau L, Bonnet A, Le Danvic C, Guyonnet B, Kiefer H, Jammes H, Schibler L. Chromatin. 2021 May 24;14(1):24. doi: 10.1186/s13072-021-00397-5. PMID: 34030709; PMCID: PMC8146655.

/bin contains all custom scripts used in the pipeline.

/Galaxy directory contains all the wrappers to use the PAQmiR approach with Galaxy. Custom scripts used in the wrapper are symbolink links to the bin folder.

[/pipeline_XX_template] are examples of projects using the PAQmiR approach. It will help you understand what is used and produced at each step of the processing. The [/pipeline_XX_template/sh-[sge|slurm]] directory contains all the scripts required to run the PAQmiR approach on a calculation server. You will need to change the relative paths into absolute paths in the scripts to be executed on the cluster (sge/slurm).

/pipeline_1_template is the first and simplest version of the pipeline. It was the version used in publications [1-4].
the main steps of the pipeline are :

• reads collapsing and mapping against reference genome using mapper.pl (from miRDeep2 suite)
• precursor/miRNA prediction using mirdeep2.pl
• creation of new precursor/miRNA dataset by merging know and predicted precursors/miRNA
• quantification and annotation of the Know/novel miRNAs using quantifier.pl (from miRDeep suite)
• post processing to remove redundancy between miRNAs
  More informations/descriptions of the pipeline can be found in the /pipeline_1_template/documentation folder.

Comment : the shell scripts provided are set up for a cluster using a sge scheduler

/pipeline_2_template This version is almost identical to template 1. The differences are as follows:

• all the pipeline parameters are defined in a config.txt file
• the samples to be analyzed must be described in a samples.txt file.
• the files containing the reads must be in fastq.gz format instead of fastq
• mapper.pl is replaced by the use of fastx_collapser + bowtie
• slurm replaces sge

the main steps of the pipeline are :

1. FASQC and multiQC
2. reads collapsing (fastx_collapser)
3. reads mapping against reference genome using bowtie
4. precursor/miRNA prediction using mirdeep2.pl and create new dataset and creation of new precursor/miRNA dataset by merging know and predicted precursors/miRNA
5. quantification and annotation of the Know/novel miRNAs using quantifier.pl (from miRDeep suite)
6. post processing to remove redundancy between miRNAs
   More informations/descriptions of the pipeline can be found in the /pipeline_2_template/documentation folder.

Comment : the shell scripts provided are set up for a cluster using a slurm scheduler and preconfigured for Genotoul Bioinformatics Facility

/pipeline_3_template is the second version of the pipeline. It is the version actually used in the majority of current projects. It was the version used in publications [5-6].
the major additions to the pipeline_1 are :

• IsomiR analysis: creation of a count table of all the miRNA IsomiRs quantified by the miRDeep2 quantifier module.
• Generic sncRNA analysis: exploitation of all unique sequences (miRNA or not):
  • creation of a general counting matrix of all the unique sequences
  • annotation of sequences against reference databases
  • merge with the results of the miRNA analysis
    More informations/descriptions of the pipeline can be found in the /pipeline_3_template/documentation folder

Comment : the shell scripts provided are set up for a cluster using a slurm scheduler