#!/usr/bin/env python """ Runs Bowtie on single-end or paired-end data. For use with Bowtie v. 0.12.3 usage: bowtie_wrapper.py [options] -t, --threads=t: The number of threads to run -i, --input1=i: The (forward or single-end) reads file in Sanger FASTQ format -I, --input2=I: The reverse reads file in Sanger FASTQ format -o, --output=o: The output file -4, --dataType=4: The type of data (SOLiD or Solexa) -2, --paired=2: Whether the data is single- or paired-end -g, --genomeSource=g: The type of reference provided -r, --ref=r: The reference genome to use or index -s, --skip=s: Skip the first n reads -a, --alignLimit=a: Only align the first n reads -T, --trimH=T: Trim n bases from high-quality (left) end of each read before alignment -L, --trimL=L: Trim n bases from low-quality (right) end of each read before alignment -m, --mismatchSeed=m: Maximum number of mismatches permitted in the seed -M, --mismatchQual=M: Maximum permitted total of quality values at mismatched read positions -l, --seedLen=l: Seed length -n, --rounding=n: Whether or not to round to the nearest 10 and saturating at 30 -P, --maqSoapAlign=P: Choose MAQ- or SOAP-like alignment policy -w, --tryHard=: Whether or not to try as hard as possible to find valid alignments when they exist -v, --valAlign=v: Report up to n valid arguments per read -V, --allValAligns=V: Whether or not to report all valid alignments per read -G, --suppressAlign=G: Suppress all alignments for a read if more than n reportable alignments exist -b, --best=b: Whether or not to make Bowtie guarantee that reported singleton alignments are 'best' in terms of stratum and in terms of the quality values at the mismatched positions -B, --maxBacktracks=B: Maximum number of backtracks permitted when aligning a read -R, --strata=R: Whether or not to report only those alignments that fall in the best stratum if many valid alignments exist and are reportable -j, --minInsert=j: Minimum insert size for valid paired-end alignments -J, --maxInsert=J: Maximum insert size for valid paired-end alignments -O, --mateOrient=O: The upstream/downstream mate orientation for valid paired-end alignment against the forward reference strand -A, --maxAlignAttempt=A: Maximum number of attempts Bowtie will make to match an alignment for one mate with an alignment for the opposite mate -f, --forwardAlign=f: Whether or not to attempt to align the forward reference strand -E, --reverseAlign=E: Whether or not to attempt to align the reverse-complement reference strand -F, --offrate=F: Override the offrate of the index to n -8, --snpphred=8: SNP penalty on Phred scale -6, --snpfrac=6: Fraction of sites expected to be SNP sites -7, --keepends=7: Keep extreme-end nucleotides and qualities -S, --seed=S: Seed for pseudo-random number generator -C, --params=C: Whether to use default or specified parameters -u, --iautoB=u: Automatic or specified behavior -K, --ipacked=K: Whether or not to use a packed representation for DNA strings -Q, --ibmax=Q: Maximum number of suffixes allowed in a block -Y, --ibmaxdivn=Y: Maximum number of suffixes allowed in a block as a fraction of the length of the reference -D, --idcv=D: The period for the difference-cover sample -U, --inodc=U: Whether or not to disable the use of the difference-cover sample -y, --inoref=y: Whether or not to build the part of the reference index used only in paired-end alignment -z, --ioffrate=z: How many rows get marked during annotation of some or all of the Burrows-Wheeler rows -W, --iftab=W: The size of the lookup table used to calculate an initial Burrows-Wheeler range with respect to the first n characters of the query -X, --intoa=X: Whether or not to convert Ns in the reference sequence to As -N, --iendian=N: Endianness to use when serializing integers to the index file -Z, --iseed=Z: Seed for the pseudorandom number generator -c, --icutoff=c: Number of first bases of the reference sequence to index -x, --indexSettings=x: Whether or not indexing options are to be set -H, --suppressHeader=H: Suppress header --do_not_build_index: Flag to specify that provided file is already indexed and to just use 'as is' """ import optparse, os, shutil, subprocess, sys, tempfile def stop_err( msg ): sys.stderr.write( '%s\n' % msg ) sys.exit() def __main__(): #Parse Command Line parser = optparse.OptionParser() parser.add_option( '-t', '--threads', dest='threads', help='The number of threads to run' ) parser.add_option( '-4', '--dataType', dest='dataType', help='The type of data (SOLiD or Solexa)' ) parser.add_option( '-i', '--input1', dest='input1', help='The (forward or single-end) reads file in Sanger FASTQ format' ) parser.add_option( '-I', '--input2', dest='input2', help='The reverse reads file in Sanger FASTQ format' ) parser.add_option( '-o', '--output', dest='output', help='The output file' ) parser.add_option( '-2', '--paired', dest='paired', help='Whether the data is single- or paired-end' ) parser.add_option( '-g', '--genomeSource', dest='genomeSource', help='The type of reference provided' ) parser.add_option( '-r', '--ref', dest='ref', help='The reference genome to use or index' ) parser.add_option( '-s', '--skip', dest='skip', help='Skip the first n reads' ) parser.add_option( '-a', '--alignLimit', dest='alignLimit', help='Only align the first n reads' ) parser.add_option( '-T', '--trimH', dest='trimH', help='Trim n bases from high-quality (left) end of each read before alignment' ) parser.add_option( '-L', '--trimL', dest='trimL', help='Trim n bases from low-quality (right) end of each read before alignment' ) parser.add_option( '-m', '--mismatchSeed', dest='mismatchSeed', help='Maximum number of mismatches permitted in the seed' ) parser.add_option( '-M', '--mismatchQual', dest='mismatchQual', help='Maximum permitted total of quality values at mismatched read positions' ) parser.add_option( '-l', '--seedLen', dest='seedLen', help='Seed length' ) parser.add_option( '-n', '--rounding', dest='rounding', help='Whether or not to round to the nearest 10 and saturating at 30' ) parser.add_option( '-P', '--maqSoapAlign', dest='maqSoapAlign', help='Choose MAQ- or SOAP-like alignment policy' ) parser.add_option( '-w', '--tryHard', dest='tryHard', help='Whether or not to try as hard as possible to find valid alignments when they exist' ) parser.add_option( '-v', '--valAlign', dest='valAlign', help='Report up to n valid arguments per read' ) parser.add_option( '-V', '--allValAligns', dest='allValAligns', help='Whether or not to report all valid alignments per read' ) parser.add_option( '-G', '--suppressAlign', dest='suppressAlign', help='Suppress all alignments for a read if more than n reportable alignments exist' ) parser.add_option( '-b', '--best', dest='best', help="Whether or not to make Bowtie guarantee that reported singleton alignments are 'best' in terms of stratum and in terms of the quality values at the mismatched positions" ) parser.add_option( '-B', '--maxBacktracks', dest='maxBacktracks', help='Maximum number of backtracks permitted when aligning a read' ) parser.add_option( '-R', '--strata', dest='strata', help='Whether or not to report only those alignments that fall in the best stratum if many valid alignments exist and are reportable' ) parser.add_option( '-j', '--minInsert', dest='minInsert', help='Minimum insert size for valid paired-end alignments' ) parser.add_option( '-J', '--maxInsert', dest='maxInsert', help='Maximum insert size for valid paired-end alignments' ) parser.add_option( '-O', '--mateOrient', dest='mateOrient', help='The upstream/downstream mate orientation for valid paired-end alignment against the forward reference strand' ) parser.add_option( '-A', '--maxAlignAttempt', dest='maxAlignAttempt', help='Maximum number of attempts Bowtie will make to match an alignment for one mate with an alignment for the opposite mate' ) parser.add_option( '-f', '--forwardAlign', dest='forwardAlign', help='Whether or not to attempt to align the forward reference strand' ) parser.add_option( '-E', '--reverseAlign', dest='reverseAlign', help='Whether or not to attempt to align the reverse-complement reference strand' ) parser.add_option( '-F', '--offrate', dest='offrate', help='Override the offrate of the index to n' ) parser.add_option( '-S', '--seed', dest='seed', help='Seed for pseudo-random number generator' ) parser.add_option( '-8', '--snpphred', dest='snpphred', help='SNP penalty on Phred scale' ) parser.add_option( '-6', '--snpfrac', dest='snpfrac', help='Fraction of sites expected to be SNP sites' ) parser.add_option( '-7', '--keepends', dest='keepends', help='Keep extreme-end nucleotides and qualities' ) parser.add_option( '-C', '--params', dest='params', help='Whether to use default or specified parameters' ) parser.add_option( '-u', '--iautoB', dest='iautoB', help='Automatic or specified behavior' ) parser.add_option( '-K', '--ipacked', dest='ipacked', help='Whether or not to use a packed representation for DNA strings' ) parser.add_option( '-Q', '--ibmax', dest='ibmax', help='Maximum number of suffixes allowed in a block' ) parser.add_option( '-Y', '--ibmaxdivn', dest='ibmaxdivn', help='Maximum number of suffixes allowed in a block as a fraction of the length of the reference' ) parser.add_option( '-D', '--idcv', dest='idcv', help='The period for the difference-cover sample' ) parser.add_option( '-U', '--inodc', dest='inodc', help='Whether or not to disable the use of the difference-cover sample' ) parser.add_option( '-y', '--inoref', dest='inoref', help='Whether or not to build the part of the reference index used only in paired-end alignment' ) parser.add_option( '-z', '--ioffrate', dest='ioffrate', help='How many rows get marked during annotation of some or all of the Burrows-Wheeler rows' ) parser.add_option( '-W', '--iftab', dest='iftab', help='The size of the lookup table used to calculate an initial Burrows-Wheeler range with respect to the first n characters of the query' ) parser.add_option( '-X', '--intoa', dest='intoa', help='Whether or not to convert Ns in the reference sequence to As' ) parser.add_option( '-N', '--iendian', dest='iendian', help='Endianness to use when serializing integers to the index file' ) parser.add_option( '-Z', '--iseed', dest='iseed', help='Seed for the pseudorandom number generator' ) parser.add_option( '-c', '--icutoff', dest='icutoff', help='Number of first bases of the reference sequence to index' ) parser.add_option( '-x', '--indexSettings', dest='index_settings', help='Whether or not indexing options are to be set' ) parser.add_option( '-H', '--suppressHeader', dest='suppressHeader', help='Suppress header' ) parser.add_option( '--do_not_build_index', dest='do_not_build_index', action="store_true", default=False, help='Flag to specify that provided file is already indexed, use as is' ) (options, args) = parser.parse_args() stdout = '' # make temp directory for placement of indices and copy reference file there if necessary tmp_index_dir = tempfile.mkdtemp() # get type of data (solid or solexa) if options.dataType == 'solid': colorspace = '-C' else: colorspace = '' # index if necessary if options.genomeSource == 'history' and not options.do_not_build_index: # set up commands if options.index_settings =='indexPreSet': indexing_cmds = '%s' % colorspace else: try: if options.iautoB != 'None' and options.iautoB == 'set': iautoB = '--noauto' else: iautoB = '' if options. ipacked != 'None' and options.ipacked == 'packed': ipacked = '--packed' else: ipacked = '' if options.ibmax != 'None' and int( options.ibmax ) >= 1: ibmax = '--bmax %s' % options.ibmax else: ibmax = '' if options.ibmaxdivn != 'None' and int( options.ibmaxdivn ) >= 0: ibmaxdivn = '--bmaxdivn %s' % options.ibmaxdivn else: ibmaxdivn = '' if options.idcv != 'None' and int( options.idcv ) > 0: idcv = '--dcv %s' % options.idcv else: idcv = '' if options.inodc != 'None' and options.inodc == 'nodc': inodc = '--nodc' else: inodc = '' if options.inoref != 'None' and options.inoref == 'noref': inoref = '--noref' else: inoref = '' if options.iftab != 'None' and int( options.iftab ) >= 0: iftab = '--ftabchars %s' % options.iftab else: iftab = '' if options.intoa != 'None' and options.intoa == 'yes': intoa = '--ntoa' else: intoa = '' if options.iendian != 'None' and options.iendian == 'big': iendian = '--big' else: iendian = '--little' if options.iseed != 'None' and int( options.iseed ) > 0: iseed = '--seed %s' % options.iseed else: iseed = '' if options.icutoff != 'None' and int( options.icutoff ) > 0: icutoff = '--cutoff %s' % options.icutoff else: icutoff = '' indexing_cmds = '%s %s %s %s %s %s %s --offrate %s %s %s %s %s %s %s' % \ ( iautoB, ipacked, ibmax, ibmaxdivn, idcv, inodc, inoref, options.ioffrate, iftab, intoa, iendian, iseed, icutoff, colorspace ) except ValueError, e: # clean up temp dir if os.path.exists( tmp_index_dir ): shutil.rmtree( tmp_index_dir ) stop_err( "Something is wrong with the indexing parameters and the indexing and alignment could not be run. Make sure you don't have any non-numeric values where they should be numeric.\n" + str( e ) ) ref_file = tempfile.NamedTemporaryFile( dir=tmp_index_dir ) ref_file_name = ref_file.name ref_file.close() os.symlink( options.ref, ref_file_name ) cmd1 = 'bowtie-build %s -f %s %s' % ( indexing_cmds, ref_file_name, ref_file_name ) try: tmp = tempfile.NamedTemporaryFile( dir=tmp_index_dir ).name tmp_stderr = open( tmp, 'wb' ) proc = subprocess.Popen( args=cmd1, shell=True, cwd=tmp_index_dir, stderr=tmp_stderr.fileno() ) returncode = proc.wait() tmp_stderr.close() # get stderr, allowing for case where it's very large tmp_stderr = open( tmp, 'rb' ) stderr = '' buffsize = 1048576 try: while True: stderr += tmp_stderr.read( buffsize ) if not stderr or len( stderr ) % buffsize != 0: break except OverflowError: pass tmp_stderr.close() if returncode != 0: raise Exception, stderr except Exception, e: # clean up temp dir if os.path.exists( tmp_index_dir ): shutil.rmtree( tmp_index_dir ) stop_err( 'Error indexing reference sequence\n' + str( e ) ) stdout += 'File indexed. ' else: ref_file_name = options.ref # set up aligning and generate aligning command options # automatically set threads in both cases if options.suppressHeader == 'true': suppressHeader = '--sam-nohead' else: suppressHeader = '' if options.maxInsert != 'None' and int( options.maxInsert ) > 0: maxInsert = '-X %s' % options.maxInsert else: maxInsert = '' if options.mateOrient != 'None': mateOrient = '--%s' % options.mateOrient else: mateOrient = '' if options.params == 'preSet': aligning_cmds = '%s %s -p %s -S %s -q %s ' % \ ( maxInsert, mateOrient, options.threads, suppressHeader, colorspace ) else: try: if options.skip != 'None' and int( options.skip ) > 0: skip = '-s %s' % options.skip else: skip = '' if options.alignLimit != 'None' and int( options.alignLimit ) >= 0: alignLimit = '-u %s' % options.alignLimit else: alignLimit = '' if options.trimH != 'None' and int( options.trimH ) > 0: trimH = '-5 %s' % options.trimH else: trimH = '' if options.trimL != 'None' and int( options.trimL ) > 0: trimL = '-3 %s' % options.trimL else: trimL = '' if options.mismatchSeed != 'None' and (options.mismatchSeed == '0' or options.mismatchSeed == '1' \ or options.mismatchSeed == '2' or options.mismatchSeed == '3'): mismatchSeed = '-n %s' % options.mismatchSeed else: mismatchSeed = '' if options.mismatchQual != 'None' and int( options.mismatchQual ) >= 0: mismatchQual = '-e %s' % options.mismatchQual else: mismatchQual = '' if options.seedLen != 'None' and int( options.seedLen ) >= 5: seedLen = '-l %s' % options.seedLen else: seedLen = '' if options.rounding == 'noRound': rounding = '--nomaqround' else: rounding = '' if options.maqSoapAlign != '-1' and int( options.maqSoapAlign ) >= 0: maqSoapAlign = '-v %s' % options.maqSoapAlign else: maqSoapAlign = '' if options.minInsert != 'None' and int( options.minInsert ) > 0: minInsert = '-I %s' % options.minInsert else: minInsert = '' if options.maxAlignAttempt != 'None' and int( options.maxAlignAttempt ) >= 0: maxAlignAttempt = '--pairtries %s' % options.maxAlignAttempt else: maxAlignAttempt = '' if options.forwardAlign == 'noForward': forwardAlign = '--nofw' else: forwardAlign = '' if options.reverseAlign == 'noReverse': reverseAlign = '--norc' else: reverseAlign = '' if options.maxBacktracks != 'None' and int( options.maxBacktracks ) > 0 and \ ( options.mismatchSeed == '2' or options.mismatchSeed == '3' ): maxBacktracks = '--maxbts %s' % options.maxBacktracks else: maxBacktracks = '' if options.tryHard == 'doTryHard': tryHard = '-y' else: tryHard = '' if options.valAlign != 'None' and int( options.valAlign ) >= 0: valAlign = '-k %s' % options.valAlign else: valAlign = '' if options.allValAligns == 'doAllValAligns': allValAligns = '-a' else: allValAligns = '' if options.suppressAlign != 'None' and int( options.suppressAlign ) >= 0: suppressAlign = '-m %s' % options.suppressAlign else: suppressAlign = '' if options.best == 'doBest': best = '--best' else: best = '' if options.strata == 'doStrata': strata = '--strata' else: strata = '' if options.offrate != 'None' and int( options.offrate ) >= 0: offrate = '-o %s' % options.offrate else: offrate = '' if options.seed != 'None' and int( options.seed ) >= 0: seed = '--seed %s' % options.seed else: seed = '' if options.snpphred != 'None' and int( options.snpphred ) >= 0: snpphred = '--snpphred %s' % options.snpphred else: snpphred = '' if options.snpfrac != 'None' and float( options.snpfrac ) >= 0: snpfrac = '--snpfrac %s' % options.snpfrac else: snpfrac = '' if options.keepends != 'None' and options.keepends == 'doKeepends': keepends = '--col-keepends' else: keepends = '' aligning_cmds = '%s %s %s %s %s %s %s %s %s %s %s %s %s %s %s %s %s ' \ '%s %s %s %s %s %s %s %s %s %s %s -p %s -S %s -q' % \ ( skip, alignLimit, trimH, trimL, mismatchSeed, mismatchQual, seedLen, rounding, maqSoapAlign, minInsert, maxInsert, mateOrient, maxAlignAttempt, forwardAlign, reverseAlign, maxBacktracks, tryHard, valAlign, allValAligns, suppressAlign, best, strata, offrate, seed, colorspace, snpphred, snpfrac, keepends, options.threads, suppressHeader ) except ValueError, e: # clean up temp dir if os.path.exists( tmp_index_dir ): shutil.rmtree( tmp_index_dir ) stop_err( 'Something is wrong with the alignment parameters and the alignment could not be run\n' + str( e ) ) try: # have to nest try-except in try-finally to handle 2.4 try: # prepare actual aligning commands if options.paired == 'paired': cmd2 = 'bowtie %s %s -1 %s -2 %s > %s' % ( aligning_cmds, ref_file_name, options.input1, options.input2, options.output ) else: cmd2 = 'bowtie %s %s %s > %s' % ( aligning_cmds, ref_file_name, options.input1, options.output ) # align tmp = tempfile.NamedTemporaryFile( dir=tmp_index_dir ).name tmp_stderr = open( tmp, 'wb' ) proc = subprocess.Popen( args=cmd2, shell=True, cwd=tmp_index_dir, stderr=tmp_stderr.fileno() ) returncode = proc.wait() tmp_stderr.close() # get stderr, allowing for case where it's very large tmp_stderr = open( tmp, 'rb' ) stderr = '' buffsize = 1048576 try: while True: stderr += tmp_stderr.read( buffsize ) if not stderr or len( stderr ) % buffsize != 0: break except OverflowError: pass tmp_stderr.close() if returncode != 0: raise Exception, stderr # check that there are results in the output file if os.path.getsize( options.output ) == 0: raise Exception, 'The output file is empty, there may be an error with your input file or settings.' + '\nextra: ' + str(extra) except Exception, e: stop_err( 'Error aligning sequence. ' + str( e ) ) finally: # clean up temp dir if os.path.exists( tmp_index_dir ): shutil.rmtree( tmp_index_dir ) stdout += 'Sequence file aligned.\n' sys.stdout.write( stdout ) if __name__=="__main__": __main__()