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miRkwood-RNA/miRkwood
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SYNOPSIS miRkwood is an application that allows for the fast and easy identification of microRNAs. It is specifically designed for plant microRNAs. INSTALL See file INSTALL.md. USAGE miRkwood comes in two distinct pipelines, according to the input data type. -mirkwood.pl (abinitio pipeline): scans a genomic sequence and finds all potential microRNA precursors. Input: a FASTA file. -mirkwood-bed.pl (smallRNAseq pipeline): analyses small RNA deep sequencing data and find all potential microRNAs. Input : a BED file. OPTIONS -mirkwood.pl: perl -I/{miRkwood_path}/cgi-bin/lib/ mirkwood.pl [options] Mandatory options: --input Path to the fasta file. --output Output directory. If non existing it will be created. The directory must be empty. Additional options: --both-strands Scan both strands. --species-mask Mask coding regions against the given organism. --shuffles Compute thermodynamic stability (shuffled sequences). --filter-mfei Select only sequences with MFEI < -0.6. --filter-rrna Filter out ribosomal RNAs (using RNAmmer). --filter-trna Filter out tRNAs (using tRNAscan-SE). --align Flag conserved mature miRNAs (alignment with miRBase + miRdup). --varna Allow the structure generation using Varna. --help Print a brief help message and exits. --man Prints the manual page and exits. -mirkwood-bed.pl: perl -I/{miRkwood_path}/cgi-bin/lib/ mirkwood-bed.pl [options] Mandatory options: --input Path to the BED file (created with our script mirkwood-bam2bed.pl). --genome Path to the genome (fasta format). --output Output directory. If non existing it will be created. The directory must be empty. Additional options: --shuffles Compute thermodynamic stability (shuffled sequences). --align Flag conserved mature miRNAs (alignment with miRBase + miRdup). --no-filter-mfei Don't filter out sequences with MFEI >= -0.6. Default : only keep sequences with MFEI < -0.6. --mirbase If you have a gff file containing known miRNAs for this assembly, use this option to give the path to this file. --gff List of annotation files (gff or gff3 format). Reads matching with an element of these files will be filtered out. For instance you can filter out CDS by providing a suitable GFF file. --no-filter-bad-hairpins By default the candidates with a quality score of 0 and no conservation are discarded from results and are stored in a BED file. Use this option to keep all results. --min-repeats Minimum number of positions for each read to be kept. Default : 0. --max-repeats Maximum number of positions for each read to be kept. Default : 5 (reads that map at more than 5 positions are filtered out). --varna Allow the structure generation using Varna. --help Print a brief help message and exits. --man Prints the manual page and exits. OUTPUT For both pipelines: alignments : folder containing all alignments files (only if option --align is on). images: folder containing images created by VARNA (only if option --varna is on). results: folder containing all results files, in several formats (csv, fa, gff, html and txt). sequences: folder containing sequences for each candidate in fasta and dotbracket format, alternatives sequences if they exist and optimal structure if it is different from the stemloop structure. YML: folder containing all candidates data in YAML format. basic_candidates.yml: contains a summary of all candidates with basic informations (this file is needed to create the results files). log.log: log file (hey, what did you expect?) run_options.cfg: config file with the chosen options. ab initio pipeline only: masks: folder containing results of BlastX, rnammer and tRNAscan-SE. input_sequences.fas: your sequences. smallRNAseq pipeline only: read_clouds: folder containing all text files for the candidates read clouds. bed_sizes.txt: tabulated file with the number of reads in each BED file. summary.txt: contains a summary of your options and of results. Depending on the options you chose for your job you may find some of the following files: your_bed_your_GFF.tar.gz: a compressed BED containing all reads matching to features from your GFF file, for each GFF file that you provided. your_bed_multimapped.tar.gz: a compressed BED containing all reads from your input BED file mapping at less than --min-repeats positions or more than --max-repeats positions. your_bed_miRNAs.tar.gz: a compressed BED containing all reads from your input BED file corresponding to miRNAs present in miRBase. your_bed_orphan_clusters.tar.gz: a compressed BED containing all reads from your input BED file that fall into a peak but that don't correspond to a valid miRNA candidate. your_bed_orphan_hairpins.tar.gz: a compressed BED containing all candidates with a quality score of 0 and no conservation. By default these candidates are excluded from final results, but you can change this behaviour with flag option --no-filter-bad-hairpins. your_bed_filtered.bed: a BED containing all reads from your input BED file that have not been filtered out in one of the previous categories.
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