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reAminator is a collection of command-line scripts to process long-read (200+ bp), high-throughput bisulfite genomic sequencing. reAminator comprises 6 Python scripts: barcode.py -- label reads by barcode bsAlign.py -- align reads to a library of references bsDraw.py -- extract aligned data from bsAlign.py output, filtering for length and percent deamination meMapper.py -- Python wrapper to call MethylMapper from bsDraw.py meTools.py -- search for methylation patterns of interest gouache.py -- draw PNG images of methylation patterns REQUIREMENTS reAminator requires Python 2.7 with the BioPython module, a local installation of BLAST+, and FASTools. These may be obtained at http://biopython.org/wiki/Main_Page ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST/ http://genome.ufl.edu/rivalab/fastools/ Optional dependencies are MethylMapper and the Python Imaging LIbrary: http://www.pythonware.com/products/pil/ Input files for barcode.py, bsAlign.py, and bsDraw.py must be in FASTA format. INSTALLATION Open the file "reAminator.cfg" in a text editor and update the paths to local BLAST+ and FASTools. Optional: update path to MethylMapper, and set whether Python Imaging Library is present (for gouache.py). USAGE usage: barcode.py [-h] [-d DIRECTORY] [-n] [-p] reads codes positional arguments: reads FASTA-format file of 454 reads codes FASTA-format file of barcodes optional arguments: -h, --help show this help message and exit -d DIRECTORY, --directory DIRECTORY output file path -n, --names track barcodes by ID (default by sequence) -p, --perfect no mismatches usage: bsAlign.py [-h] [-Save SAVE] [-mask] [-pair PAIR] [-1] [-2] seqs refs positional arguments: seqs FASTA-format file of read sequences refs FASTA-format file of reference sequences optional arguments: -h, --help show this help message and exit -Save SAVE output file name -mask enable lowercase masking in references -pair PAIR 2nd FASTA-format file for paired-end sequencing -1, --a1_or_b1 convert BS-read G to A, i.e.seq. primer = a1 or b1 -2, --a2_or_b2 convert BS-read C to T, i.e.seq. primer = a2 or b2 usage: bsDraw.py [-h] [-dest DEST] [-codes [CODES [CODES ...]]] [-refs [REFS [REFS ...]]] [-strand STRAND] [-bisulfite BISULFITE] [-length LENGTH] [-uniques] [-Sites SITES] [-TSS] [-Weights WEIGHTS] [-gouache] [-window WINDOW] [alignments [alignments ...]] positional arguments: alignments bsBlast output table file/s optional arguments: -h, --help show this help message and exit -dest DEST output directory -codes [CODES [CODES ...]] barcode seqs (leave blank for all) -refs [REFS [REFS ...]] ref. IDs (leave blank for all) -strand STRAND strands ("a" = C to T, "b" = G to A) -bisulfite BISULFITE minim. percent deaminated (0-100) -length LENGTH minim. bp length (default) or percent -uniques use only unique non-deamination patterns -Sites SITES enter alternate methylation dictionary (e.g. CG=1,CC=1) NOTE: not compatible with MethylMapper -TSS read TSS distance after "+" symbol in ref. id (e.g. >YFG+400) -Weights WEIGHTS ratio of CG to GC weight for MethylMapper (e.g. 50,50) -gouache make gouache .png files -window WINDOW basepair width of gouache .png files (or zero for automatic) EXAMPLE OF USE To label read sequences in EXAMPLE/reads.fa with barcodes from EXAMPLE/barcodes.fa, type at the command line (omit the prompt "$ "): $ python barcode.py EXAMPLE/reads.fa EXAMPLE/barcodes.fa To align the barcoded reads to sequences from EXAMPLE/references.fa: $ python bsAlign.py EXAMPLE/reads.barcodes.fa EXAMPLE/references.fa To extract all FASTA alignment files from the database EXAMPLE/reads.references.db: $ python bsDraw.py EXAMPLE/reads.references.db To extract all FASTA alignment files, and produce gouache.py PNG images: $ python bsDraw.py EXAMPLE/reads.references.db -g (c) 2013, Russell Darst, University of Florida
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