view tools/picard/picard_wrapper.py @ 1:cdcb0ce84a1b

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author xuebing
date Fri, 09 Mar 2012 19:45:15 -0500
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#!/usr/bin/env python
"""
Originally written by Kelly Vincent
pretty output and additional picard wrappers by Ross Lazarus for rgenetics
Runs all available wrapped Picard tools.
usage: picard_wrapper.py [options]
code Ross wrote licensed under the LGPL
see http://www.gnu.org/copyleft/lesser.html
"""

import optparse, os, sys, subprocess, tempfile, shutil, time, logging

galhtmlprefix = """<?xml version="1.0" encoding="utf-8" ?>
<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">
<head>
<meta http-equiv="Content-Type" content="text/html; charset=utf-8" />
<meta name="generator" content="Galaxy %s tool output - see http://getgalaxy.org/" />
<title></title>
<link rel="stylesheet" href="/static/style/base.css" type="text/css" />
</head>
<body>
<div class="document">
"""
galhtmlattr = """Galaxy tool %s run at %s</b><br/>"""
galhtmlpostfix = """</div></body></html>\n"""


def stop_err( msg ):
    sys.stderr.write( '%s\n' % msg )
    sys.exit()
    

def timenow():
    """return current time as a string
    """
    return time.strftime('%d/%m/%Y %H:%M:%S', time.localtime(time.time()))


class PicardBase():
    """
    simple base class with some utilities for Picard
    adapted and merged with Kelly Vincent's code april 2011 Ross
    lots of changes...
    """
    
    def __init__(self, opts=None,arg0=None):
        """ common stuff needed at init for a picard tool
        """
        assert opts <> None, 'PicardBase needs opts at init'
        self.opts = opts
        if self.opts.outdir == None:
             self.opts.outdir = os.getcwd() # fixmate has no html file eg so use temp dir
        assert self.opts.outdir <> None,'## PicardBase needs a temp directory if no output directory passed in'
        self.picname = self.baseName(opts.jar)
        if self.picname.startswith('picard'):
            self.picname = opts.picard_cmd # special case for some tools like replaceheader?
        self.progname = self.baseName(arg0)
        self.version = '0.002'
        self.delme = [] # list of files to destroy
        self.title = opts.title
        self.inputfile = opts.input
        try:
            os.makedirs(opts.outdir)
        except:
            pass
        try:
            os.makedirs(opts.tmpdir)
        except:
            pass
        self.log_filename = os.path.join(self.opts.outdir,'%s.log' % self.picname)
        self.metricsOut =  os.path.join(opts.outdir,'%s.metrics.txt' % self.picname)
        self.setLogging(logfname=self.log_filename)
 
    def baseName(self,name=None):
        return os.path.splitext(os.path.basename(name))[0]

    def setLogging(self,logfname="picard_wrapper.log"):
        """setup a logger
        """
        logging.basicConfig(level=logging.INFO,
                    filename=logfname,
                    filemode='a')


    def readLarge(self,fname=None):
        """ read a potentially huge file.
        """
        try:
            # get stderr, allowing for case where it's very large
            tmp = open( fname, 'rb' )
            s = ''
            buffsize = 1048576
            try:
                while True:
                    more = tmp.read( buffsize )
                    if len(more) > 0:
                        s += more
                    else:
                        break
            except OverflowError:
                pass
            tmp.close()
        except Exception, e:
            stop_err( 'Error : %s' % str( e ) )   
        return s
    
    def runCL(self,cl=None,output_dir=None):
        """ construct and run a command line
        we have galaxy's temp path as opt.temp_dir so don't really need isolation
        sometimes stdout is needed as the output - ugly hacks to deal with potentially vast artifacts
        """
        assert cl <> None, 'PicardBase runCL needs a command line as cl'
        if output_dir == None:
            output_dir = self.opts.outdir
        if type(cl) == type([]):
            cl = ' '.join(cl)
        fd,templog = tempfile.mkstemp(dir=output_dir,suffix='rgtempRun.txt')
        tlf = open(templog,'wb')
        fd,temperr = tempfile.mkstemp(dir=output_dir,suffix='rgtempErr.txt')
        tef = open(temperr,'wb')
        process = subprocess.Popen(cl, shell=True, stderr=tef, stdout=tlf, cwd=output_dir)
        rval = process.wait()
        tlf.close()
        tef.close()
        stderrs = self.readLarge(temperr)
        stdouts = self.readLarge(templog)        
        if len(stderrs) > 0:
            s = '## executing %s returned status %d and stderr: \n%s\n' % (cl,rval,stderrs)
        else:
            s = '## executing %s returned status %d and nothing on stderr\n' % (cl,rval)
        logging.info(s)
        os.unlink(templog) # always
        os.unlink(temperr) # always
        return s, stdouts # sometimes this is an output
    
    def runPic(self, jar, cl):
        """
        cl should be everything after the jar file name in the command
        """
        runme = ['java -Xmx%s' % self.opts.maxjheap]
        runme.append('-jar %s' % jar)
        runme += cl
        s,stdout = self.runCL(cl=runme, output_dir=self.opts.outdir)
        return stdout

    def samToBam(self,infile=None,outdir=None):
        """
        use samtools view to convert sam to bam
        """
        fd,tempbam = tempfile.mkstemp(dir=outdir,suffix='rgutilsTemp.bam')
        cl = ['samtools view -h -b -S -o ',tempbam,infile]
        tlog,stdouts = self.runCL(cl,outdir)
        return tlog,tempbam

    #def bamToSam(self,infile=None,outdir=None):
    #    """
    #    use samtools view to convert bam to sam
    #    """
    #    fd,tempsam = tempfile.mkstemp(dir=outdir,suffix='rgutilsTemp.sam')
    #    cl = ['samtools view -h -o ',tempsam,infile]
    #    tlog,stdouts = self.runCL(cl,outdir)
    #    return tlog,tempsam

    def sortSam(self, infile=None,outfile=None,outdir=None):
        """
        """
        print '## sortSam got infile=%s,outfile=%s,outdir=%s' % (infile,outfile,outdir)
        cl = ['samtools sort',infile,outfile]
        tlog,stdouts = self.runCL(cl,outdir)
        return tlog

    def cleanup(self):
        for fname in self.delme:
            try:
                os.unlink(fname)
            except:
                pass
                    
    def prettyPicout(self,transpose,maxrows):
        """organize picard outpouts into a report html page
        """
        res = []
        try:
            r = open(self.metricsOut,'r').readlines()
        except:
            r = []        
        if len(r) > 0:
            res.append('<b>Picard on line resources</b><ul>\n')
            res.append('<li><a href="http://picard.sourceforge.net/index.shtml">Click here for Picard Documentation</a></li>\n')
            res.append('<li><a href="http://picard.sourceforge.net/picard-metric-definitions.shtml">Click here for Picard Metrics definitions</a></li></ul><hr/>\n')
            if transpose:
                res.append('<b>Picard output (transposed to make it easier to see)</b><hr/>\n')       
            else:
                res.append('<b>Picard output</b><hr/>\n')  
            res.append('<table cellpadding="3" >\n')
            dat = []
            heads = []
            lastr = len(r) - 1
            # special case for estimate library complexity hist
            thist = False
            for i,row in enumerate(r):
                if row.strip() > '':
                    srow = row.split('\t')
                    if row.startswith('#'):
                        heads.append(row.strip()) # want strings
                    else:
                        dat.append(srow) # want lists
                    if row.startswith('## HISTOGRAM'):
                        thist = True
            if len(heads) > 0:
                hres = ['<tr class="d%d"><td colspan="2">%s</td></tr>' % (i % 2,x) for i,x in enumerate(heads)]
                res += hres
                heads = []
            if len(dat) > 0:
                if transpose and not thist:
                    tdat = map(None,*dat) # transpose an arbitrary list of lists
                    tdat = ['<tr class="d%d"><td>%s</td><td>%s&nbsp;</td></tr>\n' % ((i+len(heads)) % 2,x[0],x[1]) for i,x in enumerate(tdat)] 
                else:
                    tdat = ['\t'.join(x).strip() for x in dat] # back to strings :(
                    tdat = ['<tr class="d%d"><td colspan="2">%s</td></tr>\n' % ((i+len(heads)) % 2,x) for i,x in enumerate(tdat)]
                res += tdat
                dat = []
            res.append('</table>\n')   
        return res

    def fixPicardOutputs(self,transpose,maxloglines):
        """
        picard produces long hard to read tab header files
        make them available but present them transposed for readability
        """
        logging.shutdown()
        self.cleanup() # remove temp files stored in delme
        rstyle="""<style type="text/css">
        tr.d0 td {background-color: oldlace; color: black;}
        tr.d1 td {background-color: aliceblue; color: black;}
        </style>"""    
        res = [rstyle,]
        res.append(galhtmlprefix % self.progname)   
        res.append(galhtmlattr % (self.picname,timenow()))
        flist = [x for x in os.listdir(self.opts.outdir) if not x.startswith('.')] 
        pdflist = [x for x in flist if os.path.splitext(x)[-1].lower() == '.pdf']
        if len(pdflist) > 0: # assumes all pdfs come with thumbnail .jpgs
            for p in pdflist:
                imghref = '%s.jpg' % os.path.splitext(p)[0] # removes .pdf
                res.append('<table cellpadding="10"><tr><td>\n')
                res.append('<a href="%s"><img src="%s" title="Click image preview for a print quality PDF version" hspace="10" align="middle"></a>\n' % (p,imghref)) 
                res.append('</tr></td></table>\n')   
        if len(flist) > 0:
            res.append('<b>The following output files were created (click the filename to view/download a copy):</b><hr/>')
            res.append('<table>\n')
            for i,f in enumerate(flist):
                fn = os.path.split(f)[-1]
                res.append('<tr><td><a href="%s">%s</a></td></tr>\n' % (fn,fn))
            res.append('</table><p/>\n') 
        pres = self.prettyPicout(transpose,maxloglines)
        if len(pres) > 0:
            res += pres
        l = open(self.log_filename,'r').readlines()
        llen = len(l)
        if llen > 0: 
            res.append('<b>Picard Tool Run Log</b><hr/>\n') 
            rlog = ['<pre>',]
            if llen > maxloglines:
                n = min(50,int(maxloglines/2))
                rlog += l[:n]
                rlog.append('------------ ## %d rows deleted ## --------------\n' % (llen-maxloglines))
                rlog += l[-n:]
            else:
                rlog += l
            rlog.append('</pre>')
            if llen > maxloglines:
                rlog.append('\n<b>## WARNING - %d log lines truncated - <a href="%s">%s</a> contains entire output</b>' % (llen - maxloglines,self.log_filename,self.log_filename))
            res += rlog
        else:
            res.append("### Odd, Picard left no log file %s - must have really barfed badly?\n" % self.log_filename)
        res.append('<hr/>The freely available <a href="http://picard.sourceforge.net/command-line-overview.shtml">Picard software</a> \n') 
        res.append( 'generated all outputs reported here running as a <a href="http://getgalaxy.org">Galaxy</a> tool')   
        res.append(galhtmlpostfix) 
        outf = open(self.opts.htmlout,'w')
        outf.write(''.join(res))   
        outf.write('\n')
        outf.close()

    def makePicInterval(self,inbed=None,outf=None):
        """
        picard wants bait and target files to have the same header length as the incoming bam/sam 
        a meaningful (ie accurate) representation will fail because of this - so this hack
        it would be far better to be able to supply the original bed untouched
        """
        assert inbed <> None
        bed = open(inbed,'r').readlines()
        thead = os.path.join(self.opts.outdir,'tempSamHead.txt')
        if self.opts.datatype == 'sam':
            cl = ['samtools view -H -S',self.opts.input,'>',thead]
        else:
            cl = ['samtools view -H',self.opts.input,'>',thead]
        self.runCL(cl=cl,output_dir=self.opts.outdir)
        head = open(thead,'r').readlines()
        s = '## got %d rows of header\n' % (len(head))
        logging.info(s)
        o = open(outf,'w')
        o.write(''.join(head))
        o.write(''.join(bed))
        o.close()
        return outf

    def cleanSam(self, insam=None, newsam=None, picardErrors=[],outformat=None):
        """
        interesting problem - if paired, must remove mate pair of errors too or we have a new set of errors after cleaning - missing mate pairs!
        Do the work of removing all the error sequences
        pysam is cool
        infile = pysam.Samfile( "-", "r" )
        outfile = pysam.Samfile( "-", "w", template = infile )
        for s in infile: outfile.write(s)

        errors from ValidateSameFile.jar look like
        WARNING: Record 32, Read name SRR006041.1202260, NM tag (nucleotide differences) is missing
        ERROR: Record 33, Read name SRR006041.1042721, Empty sequence dictionary.
        ERROR: Record 33, Read name SRR006041.1042721, RG ID on SAMRecord not found in header: SRR006041

        """
        assert os.path.isfile(insam), 'rgPicardValidate cleansam needs an input sam file - cannot find %s' % insam
        assert newsam <> None, 'rgPicardValidate cleansam needs an output new sam file path'
        removeNames = [x.split(',')[1].replace(' Read name ','') for x in picardErrors if len(x.split(',')) > 2]
        remDict = dict(zip(removeNames,range(len(removeNames))))
        infile = pysam.Samfile(insam,'rb')
        info = 'found %d error sequences in picardErrors, %d unique' % (len(removeNames),len(remDict))
        if len(removeNames) > 0:
            outfile = pysam.Samfile(newsam,'wb',template=infile) # template must be an open file
            i = 0
            j = 0
            for row in infile:
                dropme = remDict.get(row.qname,None) # keep if None
                if not dropme:
                    outfile.write(row)
                    j += 1
                else: # discard
                    i += 1
            info = '%s\n%s' % (info, 'Discarded %d lines writing %d to %s from %s' % (i,j,newsam,insam))
            outfile.close()
            infile.close()
        else: # we really want a nullop or a simple pointer copy
            infile.close()
            if newsam:
                shutil.copy(insam,newsam)
        logging.info(info)
                


def __main__():
    doFix = False # tools returning htmlfile don't need this
    doTranspose = True # default
    maxloglines = 100 # default 
    #Parse Command Line
    op = optparse.OptionParser()
    # All tools
    op.add_option('-i', '--input', dest='input', help='Input SAM or BAM file' )
    op.add_option('-e', '--inputext', default=None)
    op.add_option('-o', '--output', default=None)
    op.add_option('-n', '--title', default="Pick a Picard Tool")
    op.add_option('-t', '--htmlout', default=None)
    op.add_option('-d', '--outdir', default=None)
    op.add_option('-x', '--maxjheap', default='4g')
    op.add_option('-b', '--bisulphite', default='false')
    op.add_option('-s', '--sortorder', default='query')     
    op.add_option('','--tmpdir', default='/tmp')
    op.add_option('-j','--jar',default='')    
    op.add_option('','--picard-cmd',default=None)    
    # Many tools
    op.add_option( '', '--output-format', dest='output_format', help='Output format' )
    op.add_option( '', '--bai-file', dest='bai_file', help='The path to the index file for the input bam file' )
    op.add_option( '', '--ref', dest='ref', help='Built-in reference with fasta and dict file', default=None )
    # CreateSequenceDictionary
    op.add_option( '', '--ref-file', dest='ref_file', help='Fasta to use as reference', default=None )
    op.add_option( '', '--species-name', dest='species_name', help='Species name to use in creating dict file from fasta file' )
    op.add_option( '', '--build-name', dest='build_name', help='Name of genome assembly to use in creating dict file from fasta file' )
    op.add_option( '', '--trunc-names', dest='trunc_names', help='Truncate sequence names at first whitespace from fasta file' )
    # MarkDuplicates
    op.add_option( '', '--remdups', default='true', help='Remove duplicates from output file' )
    op.add_option( '', '--optdupdist', default="100", help='Maximum pixels between two identical sequences in order to consider them optical duplicates.' )
    # CollectInsertSizeMetrics
    op.add_option('', '--taillimit', default="0")
    op.add_option('', '--histwidth', default="0")
    op.add_option('', '--minpct', default="0.01")
    # CollectAlignmentSummaryMetrics
    op.add_option('', '--maxinsert', default="20")
    op.add_option('', '--adaptors', action='append', type="string")
    # FixMateInformation and validate
    # CollectGcBiasMetrics
    op.add_option('', '--windowsize', default='100')
    op.add_option('', '--mingenomefrac', default='0.00001')    
    # AddOrReplaceReadGroups
    op.add_option( '', '--rg-opts', dest='rg_opts', help='Specify extra (optional) arguments with full, otherwise preSet' )
    op.add_option( '', '--rg-lb', dest='rg_library', help='Read Group Library' )
    op.add_option( '', '--rg-pl', dest='rg_platform', help='Read Group platform (e.g. illumina, solid)' )
    op.add_option( '', '--rg-pu', dest='rg_plat_unit', help='Read Group platform unit (eg. run barcode) ' )
    op.add_option( '', '--rg-sm', dest='rg_sample', help='Read Group sample name' )
    op.add_option( '', '--rg-id', dest='rg_id', help='Read Group ID' )
    op.add_option( '', '--rg-cn', dest='rg_seq_center', help='Read Group sequencing center name' )
    op.add_option( '', '--rg-ds', dest='rg_desc', help='Read Group description' )
    # ReorderSam
    op.add_option( '', '--allow-inc-dict-concord', dest='allow_inc_dict_concord', help='Allow incomplete dict concordance' )
    op.add_option( '', '--allow-contig-len-discord', dest='allow_contig_len_discord', help='Allow contig length discordance' )
    # ReplaceSamHeader
    op.add_option( '', '--header-file', dest='header_file', help='sam or bam file from which header will be read' )

    op.add_option('','--assumesorted', default='true') 
    op.add_option('','--readregex', default="[a-zA-Z0-9]+:[0-9]:([0-9]+):([0-9]+):([0-9]+).*")
    #estimatelibrarycomplexity
    op.add_option('','--minid', default="5")
    op.add_option('','--maxdiff', default="0.03")
    op.add_option('','--minmeanq', default="20")
    #hsmetrics
    op.add_option('','--baitbed', default=None)
    op.add_option('','--targetbed', default=None)
    #validate
    op.add_option('','--ignoreflags', action='append', type="string")
    op.add_option('','--maxerrors', default=None)
    op.add_option('','--datatype', default=None)
    op.add_option('','--bamout', default=None)
    op.add_option('','--samout', default=None)

    opts, args = op.parse_args()
    opts.sortme = opts.assumesorted == 'false'
    assert opts.input <> None
    # need to add
    # instance that does all the work
    pic = PicardBase(opts,sys.argv[0])

    tmp_dir = opts.outdir
    haveTempout = False # we use this where sam output is an option

    # set ref and dict files to use (create if necessary)
    ref_file_name = opts.ref
    if opts.ref_file <> None:
        csd = 'CreateSequenceDictionary'
        realjarpath = os.path.split(opts.jar)[0]
        jarpath = os.path.join(realjarpath,'%s.jar' % csd) # for refseq
        tmp_ref_fd, tmp_ref_name = tempfile.mkstemp( dir=opts.tmpdir , prefix = pic.picname)
        ref_file_name = '%s.fasta' % tmp_ref_name
        # build dict
        dict_file_name = '%s.dict' % tmp_ref_name
        os.symlink( opts.ref_file, ref_file_name )
        cl = ['REFERENCE=%s' % ref_file_name]
        cl.append('OUTPUT=%s' % dict_file_name)
        cl.append('URI=%s' % os.path.basename( opts.ref_file ))
        cl.append('TRUNCATE_NAMES_AT_WHITESPACE=%s' % opts.trunc_names)
        if opts.species_name:
            cl.append('SPECIES=%s' % opts.species_name)
        if opts.build_name:
            cl.append('GENOME_ASSEMBLY=%s' % opts.build_name)
        pic.delme.append(dict_file_name)
        pic.delme.append(ref_file_name)
        pic.delme.append(tmp_ref_name)
        s = pic.runPic(jarpath, cl)
        # run relevant command(s)

    # define temporary output
    # if output is sam, it must have that extension, otherwise bam will be produced
    # specify sam or bam file with extension
    if opts.output_format == 'sam':
        suff = '.sam'
    else:
        suff = ''
    tmp_fd, tempout = tempfile.mkstemp( dir=opts.tmpdir, suffix=suff )

    cl = ['VALIDATION_STRINGENCY=LENIENT',]

    if pic.picname == 'AddOrReplaceReadGroups':
        # sort order to match Galaxy's default
        cl.append('SORT_ORDER=coordinate')
        # input
        cl.append('INPUT=%s' % opts.input)
        # outputs
        cl.append('OUTPUT=%s' % tempout)
        # required read groups
        cl.append('RGLB="%s"' % opts.rg_library)
        cl.append('RGPL="%s"' % opts.rg_platform)
        cl.append('RGPU="%s"' % opts.rg_plat_unit)
        cl.append('RGSM="%s"' % opts.rg_sample)
        if opts.rg_id:
            cl.append('RGID="%s"' % opts.rg_id)
        # optional read groups
        if opts.rg_seq_center:
            cl.append('RGCN="%s"' % opts.rg_seq_center)
        if opts.rg_desc:
            cl.append('RGDS="%s"' % opts.rg_desc)
        pic.runPic(opts.jar, cl)
        haveTempout = True

    elif pic.picname == 'BamIndexStats':
        tmp_fd, tmp_name = tempfile.mkstemp( dir=tmp_dir )
        tmp_bam_name = '%s.bam' % tmp_name
        tmp_bai_name = '%s.bai' % tmp_bam_name
        os.symlink( opts.input, tmp_bam_name )
        os.symlink( opts.bai_file, tmp_bai_name )
        cl.append('INPUT=%s' % ( tmp_bam_name ))
        pic.delme.append(tmp_bam_name)
        pic.delme.append(tmp_bai_name)
        pic.delme.append(tmp_name)
        s = pic.runPic( opts.jar, cl )
        f = open(pic.metricsOut,'a')
        f.write(s) # got this on stdout from runCl
        f.write('\n')
        f.close()
        doTranspose = False # but not transposed

    elif pic.picname == 'EstimateLibraryComplexity':
        cl.append('I=%s' % opts.input)
        cl.append('O=%s' % pic.metricsOut)
        if float(opts.minid) > 0:
            cl.append('MIN_IDENTICAL_BASES=%s' % opts.minid)
        if float(opts.maxdiff) > 0.0:
            cl.append('MAX_DIFF_RATE=%s' % opts.maxdiff)
        if float(opts.minmeanq) > 0:
            cl.append('MIN_MEAN_QUALITY=%s' % opts.minmeanq)
        if opts.readregex > '':
            cl.append('READ_NAME_REGEX="%s"' % opts.readregex)
        if float(opts.optdupdist) > 0:
            cl.append('OPTICAL_DUPLICATE_PIXEL_DISTANCE=%s' % opts.optdupdist)
        pic.runPic(opts.jar,cl)

    elif pic.picname == 'CollectAlignmentSummaryMetrics':
        # Why do we do this fakefasta thing? Because we need NO fai to be available or picard barfs unless it has the same length as the input data.
        # why? Dunno Seems to work without complaining if the .bai file is AWOL....
        fakefasta = os.path.join(opts.outdir,'%s_fake.fasta' % os.path.basename(ref_file_name))
        try:
            os.symlink(ref_file_name,fakefasta)
        except:
            s = '## unable to symlink %s to %s - different devices? May need to replace with shutil.copy'
            info = s
            shutil.copy(ref_file_name,fakefasta)
        pic.delme.append(fakefasta)
        cl.append('ASSUME_SORTED=%s' % opts.assumesorted)
        adaptorseqs = ''.join([' ADAPTER_SEQUENCE=%s' % x for x in opts.adaptors])
        cl.append(adaptorseqs)
        cl.append('IS_BISULFITE_SEQUENCED=%s' % opts.bisulphite)
        cl.append('MAX_INSERT_SIZE=%s' % opts.maxinsert)
        cl.append('OUTPUT=%s' % pic.metricsOut)
        cl.append('R=%s' % fakefasta)
        cl.append('TMP_DIR=%s' % opts.tmpdir)
        if not opts.assumesorted.lower() == 'true': # we need to sort input
            fakeinput = '%s.sorted' % opts.input
            s = pic.sortSam(opts.input, fakeinput, opts.outdir)
            pic.delme.append(fakeinput)
            cl.append('INPUT=%s' % fakeinput)
        else:
            cl.append('INPUT=%s' % os.path.abspath(opts.input)) 
        pic.runPic(opts.jar,cl)
       
        
    elif pic.picname == 'CollectGcBiasMetrics':
        assert os.path.isfile(ref_file_name),'PicardGC needs a reference sequence - cannot read %s' % ref_file_name
        # sigh. Why do we do this fakefasta thing? Because we need NO fai to be available or picard barfs unless it has the same length as the input data.
        # why? Dunno 
        fakefasta = os.path.join(opts.outdir,'%s_fake.fasta' % os.path.basename(ref_file_name))
        try:
            os.symlink(ref_file_name,fakefasta)
        except:
            s = '## unable to symlink %s to %s - different devices? May need to replace with shutil.copy'
            info = s
            shutil.copy(ref_file_name,fakefasta)
        pic.delme.append(fakefasta)
        x = 'rgPicardGCBiasMetrics'
        pdfname = '%s.pdf' % x
        jpgname = '%s.jpg' % x
        tempout = os.path.join(opts.outdir,'rgPicardGCBiasMetrics.out')
        temppdf = os.path.join(opts.outdir,pdfname)
        cl.append('R=%s' % fakefasta)
        cl.append('WINDOW_SIZE=%s' % opts.windowsize)
        cl.append('MINIMUM_GENOME_FRACTION=%s' % opts.mingenomefrac)
        cl.append('INPUT=%s' % opts.input)
        cl.append('OUTPUT=%s' % tempout)
        cl.append('TMP_DIR=%s' % opts.tmpdir)
        cl.append('CHART_OUTPUT=%s' % temppdf)
        cl.append('SUMMARY_OUTPUT=%s' % pic.metricsOut)
        pic.runPic(opts.jar,cl)
        if os.path.isfile(temppdf):
            cl2 = ['convert','-resize x400',temppdf,os.path.join(opts.outdir,jpgname)] # make the jpg for fixPicardOutputs to find
            s,stdouts = pic.runCL(cl=cl2,output_dir=opts.outdir)
        else:
            s='### runGC: Unable to find pdf %s - please check the log for the causal problem\n' % temppdf
        lf = open(pic.log_filename,'a')
        lf.write(s)
        lf.write('\n')
        lf.close()
        
    elif pic.picname == 'CollectInsertSizeMetrics':
        isPDF = 'InsertSizeHist.pdf'
        pdfpath = os.path.join(opts.outdir,isPDF)
        histpdf = 'InsertSizeHist.pdf'
        cl.append('I=%s' % opts.input)
        cl.append('O=%s' % pic.metricsOut)
        cl.append('HISTOGRAM_FILE=%s' % histpdf)
        if opts.taillimit <> '0':
            cl.append('TAIL_LIMIT=%s' % opts.taillimit)
        if  opts.histwidth <> '0':
            cl.append('HISTOGRAM_WIDTH=%s' % opts.histwidth)
        if float( opts.minpct) > 0.0:
            cl.append('MINIMUM_PCT=%s' % opts.minpct)
        pic.runPic(opts.jar,cl)   
        if os.path.exists(pdfpath): # automake thumbnail - will be added to html 
            cl2 = ['mogrify', '-format jpg -resize x400 %s' % pdfpath]
            s,stdouts = pic.runCL(cl=cl2,output_dir=opts.outdir)
        else:
            s = 'Unable to find expected pdf file %s<br/>\n' % pdfpath
            s += 'This <b>always happens if single ended data was provided</b> to this tool,\n'
            s += 'so please double check that your input data really is paired-end NGS data.<br/>\n'
            s += 'If your input was paired data this may be a bug worth reporting to the galaxy-bugs list\n<br/>'
            stdouts = ''
        logging.info(s)
        if len(stdouts) > 0:
           logging.info(stdouts)
        
    elif pic.picname == 'MarkDuplicates':
        # assume sorted even if header says otherwise
        cl.append('ASSUME_SORTED=%s' % (opts.assumesorted))
        # input
        cl.append('INPUT=%s' % opts.input)
        # outputs
        cl.append('OUTPUT=%s' % opts.output) 
        cl.append('METRICS_FILE=%s' % pic.metricsOut )
        # remove or mark duplicates
        cl.append('REMOVE_DUPLICATES=%s' % opts.remdups)
        # the regular expression to be used to parse reads in incoming SAM file
        cl.append('READ_NAME_REGEX="%s"' % opts.readregex)
        # maximum offset between two duplicate clusters
        cl.append('OPTICAL_DUPLICATE_PIXEL_DISTANCE=%s' % opts.optdupdist)
        pic.runPic(opts.jar, cl)

    elif pic.picname == 'FixMateInformation':
        cl.append('I=%s' % opts.input)
        cl.append('O=%s' % tempout)
        cl.append('SORT_ORDER=%s' % opts.sortorder)
        pic.runPic(opts.jar,cl)
        haveTempout = True
        
    elif pic.picname == 'ReorderSam':
        # input
        cl.append('INPUT=%s' % opts.input)
        # output
        cl.append('OUTPUT=%s' % tempout)
        # reference
        cl.append('REFERENCE=%s' % ref_file_name)
        # incomplete dict concordance
        if opts.allow_inc_dict_concord == 'true':
            cl.append('ALLOW_INCOMPLETE_DICT_CONCORDANCE=true')
        # contig length discordance
        if opts.allow_contig_len_discord == 'true':
            cl.append('ALLOW_CONTIG_LENGTH_DISCORDANCE=true')
        pic.runPic(opts.jar, cl)
        haveTempout = True

    elif pic.picname == 'ReplaceSamHeader':
        cl.append('INPUT=%s' % opts.input)
        cl.append('OUTPUT=%s' % tempout)
        cl.append('HEADER=%s' % opts.header_file)
        pic.runPic(opts.jar, cl)
        haveTempout = True

    elif pic.picname == 'CalculateHsMetrics':
        maxloglines = 100
        baitfname = os.path.join(opts.outdir,'rgPicardHsMetrics.bait')
        targetfname = os.path.join(opts.outdir,'rgPicardHsMetrics.target')
        baitf = pic.makePicInterval(opts.baitbed,baitfname)
        if opts.targetbed == opts.baitbed: # same file sometimes
            targetf = baitf
        else:
            targetf = pic.makePicInterval(opts.targetbed,targetfname)   
        cl.append('BAIT_INTERVALS=%s' % baitf)
        cl.append('TARGET_INTERVALS=%s' % targetf)
        cl.append('INPUT=%s' % os.path.abspath(opts.input))
        cl.append('OUTPUT=%s' % pic.metricsOut)
        cl.append('TMP_DIR=%s' % opts.tmpdir)
        pic.runPic(opts.jar,cl)
           
    elif pic.picname == 'ValidateSamFile':
        import pysam
        doTranspose = False
        sortedfile = os.path.join(opts.outdir,'rgValidate.sorted')
        stf = open(pic.log_filename,'w')
        tlog = None
        if opts.datatype == 'sam': # need to work with a bam 
            tlog,tempbam = pic.samToBam(opts.input,opts.outdir)
            try:
                tlog = pic.sortSam(tempbam,sortedfile,opts.outdir)
            except:
                print '## exception on sorting sam file %s' % opts.input
        else: # is already bam
            try:
                tlog = pic.sortSam(opts.input,sortedfile,opts.outdir)
            except: # bug - [bam_sort_core] not being ignored - TODO fixme
                print '## exception on sorting bam file %s' % opts.input
        if tlog:
            print '##tlog=',tlog
            stf.write(tlog)
            stf.write('\n')
        sortedfile = '%s.bam' % sortedfile # samtools does that      
        cl.append('O=%s' % pic.metricsOut)
        cl.append('TMP_DIR=%s' % opts.tmpdir)
        cl.append('I=%s' % sortedfile)
        opts.maxerrors = '99999999'
        cl.append('MAX_OUTPUT=%s' % opts.maxerrors)
        if opts.ignoreflags[0] <> 'None': # picard error values to ignore
            igs = ['IGNORE=%s' % x for x in opts.ignoreflags if x <> 'None']
            cl.append(' '.join(igs))
        if opts.bisulphite.lower() <> 'false':
            cl.append('IS_BISULFITE_SEQUENCED=true')
        if opts.ref <> None or opts.ref_file <> None:
            cl.append('R=%s' %  ref_file_name)
        pic.runPic(opts.jar,cl)
        if opts.datatype == 'sam':
            pic.delme.append(tempbam)
        newsam = opts.output
        outformat = 'bam'
        pe = open(pic.metricsOut,'r').readlines()
        pic.cleanSam(insam=sortedfile, newsam=newsam, picardErrors=pe,outformat=outformat)
        pic.delme.append(sortedfile) # not wanted
        stf.close()
        pic.cleanup()
    else:
        print >> sys.stderr,'picard.py got an unknown tool name - %s' % pic.picname
        sys.exit(1)
    if haveTempout:
        # Some Picard tools produced a potentially intermediate bam file. 
        # Either just move to final location or create sam
        shutil.move(tempout, os.path.abspath(opts.output))

    if opts.htmlout <> None or doFix: # return a pretty html page
        pic.fixPicardOutputs(transpose=doTranspose,maxloglines=maxloglines)

if __name__=="__main__": __main__()