diff --git a/java/src/org/broadinstitute/sting/playground/indels/IndelInspectorMain.java b/java/src/org/broadinstitute/sting/playground/indels/IndelInspectorMain.java new file mode 100755 index 000000000..8070c3e26 --- /dev/null +++ b/java/src/org/broadinstitute/sting/playground/indels/IndelInspectorMain.java @@ -0,0 +1,313 @@ +package org.broadinstitute.sting.playground.indels; + +import java.io.File; +import java.util.List; +import java.util.Map; +import java.util.HashMap; + + +import javax.swing.JFileChooser; +import javax.swing.filechooser.FileNameExtensionFilter; +import edu.mit.broad.picard.cmdline.CommandLineProgram; +import edu.mit.broad.picard.cmdline.Option; +import edu.mit.broad.picard.cmdline.Usage; +import edu.mit.broad.picard.reference.ReferenceSequenceFileWalker; +import edu.mit.broad.picard.reference.ReferenceSequence; + +import net.sf.samtools.*; +import org.broadinstitute.sting.utils.GenomeLoc; + +public class IndelInspector extends CommandLineProgram { + + // Usage and parameters + @Usage(programVersion="1.0") public String USAGE = "Investigates indels called in the alignment data\n"; + @Option(shortName="I", doc="SAM or BAM file for calling",optional=true) public File INPUT_FILE; + @Option(shortName="L",doc="Genomic interval to run on, as contig[:start[-stop]]; whole genome if not specified", optional=true) public String GENOME_LOCATION; + @Option(shortName="V",doc="Verbosity level: SILENT, PILESUMMARY, ALIGNMENTS", optional=true) public String VERBOSITY_LEVEL; + @Option(doc="Output file (sam or bam) for non-indel related reads and indel reads that were not improved") public String OUT1; + @Option(doc="Output file (sam or bam) for improved (realigned) indel related reads") public String OUT2; + @Option(doc="[paranoid] If true, all reads that would be otherwise picked and processed by this tool will be saved, unmodified, into OUT1", optional=true) public Boolean CONTROL_RUN; + @Option(doc="Error counting mode: MM - mismatches only (from sam tags), MC - mismatches only doing actual mismatch count on the fly (use this if tags are incorrectly set); ERR - errors (arachne style: mm+gap lengths), MG - count mismatches and gaps as one error each") public String ERR_MODE; + @Option(doc="Maximum number of errors allowed (see ERR_MODE)") public Integer MAX_ERRS; + @Option(shortName="R", doc="Reference fasta or fasta.gz file") public File REF_FILE; + @Option(doc="Ignore reads that are longer than the specified cutoff (not a good way to do things but might be necessary because of performance issues)", optional=true) public Integer MAX_READ_LENGTH; + + /** Required main method implementation. */ + public static void main(final String[] argv) { + System.exit(new IndelInspector().instanceMain(argv)); + } + + protected int doWork() { + + int discarded_cigar_count = 0; + int discarded_long_read_count = 0; + + ReferenceSequenceFileWalker reference = new ReferenceSequenceFileWalker( + REF_FILE + ); + + if ( reference.getSequenceDictionary() == null ) { + System.out.println("No reference sequence dictionary found. Abort."); + } + + GenomeLoc.setupRefContigOrdering(reference.getSequenceDictionary()); + GenomeLoc location = null; + if ( GENOME_LOCATION != null ) { + location = GenomeLoc.parseGenomeLoc(GENOME_LOCATION); + } + + if ( ! ERR_MODE.equals("MM") && ! ERR_MODE.equals("MG") && ! ERR_MODE.equals("ERR") && ! ERR_MODE.equals("MC") ) { + System.out.println("Unknown value specified for ERR_MODE: "+ERR_MODE); + return 1; + } + + final SAMFileReader samReader = new SAMFileReader(getInputFile(INPUT_FILE,"/broad/1KG/")); + samReader.setValidationStringency(SAMFileReader.ValidationStringency.SILENT); + + // setContigOrdering(samReader); + + + if ( MAX_READ_LENGTH == null ) MAX_READ_LENGTH = 1000000000; + + ReferenceSequence contig_seq = null; + + IndelRecordPileCollector col = null; + PassThroughWriter ptWriter = new PassThroughWriter(OUT1,samReader.getFileHeader()); + PileBuilder pileBuilder = null; + if ( CONTROL_RUN == null ) CONTROL_RUN=false; + if ( ! CONTROL_RUN ) pileBuilder = new PileBuilder(OUT2,samReader.getFileHeader(),ptWriter); + + try { + if ( CONTROL_RUN ) col = new IndelRecordPileCollector(ptWriter, new DiscardingPileReceiver() ); + else col = new IndelRecordPileCollector(ptWriter, pileBuilder ); + } catch(Exception e) { System.err.println(e.getMessage()); } + if ( col == null ) return 1; + + col.setControlRun(CONTROL_RUN); + + if ( ! CONTROL_RUN ) { + if ( VERBOSITY_LEVEL == null ) VERBOSITY_LEVEL = new String("SILENT"); + if ( VERBOSITY_LEVEL.toUpperCase().equals("SILENT")) pileBuilder.setVerbosity(PileBuilder.SILENT); + else if ( VERBOSITY_LEVEL.toUpperCase().equals("PILESUMMARY") ) pileBuilder.setVerbosity(PileBuilder.PILESUMMARY); + else if ( VERBOSITY_LEVEL.toUpperCase().equals("ALIGNMENTS") ) pileBuilder.setVerbosity(PileBuilder.ALIGNMENTS); + else { + System.out.println("Unrecognized VERBOSITY_LEVEL setting."); + return 1; + } + } + + String cur_contig = null; + int counter = 0; + + for ( SAMRecord r : samReader ) { + + if ( r.getReadUnmappedFlag() ) continue; + if ( r.getReferenceName() != cur_contig) { + cur_contig = r.getReferenceName(); + System.out.println("Contig "+cur_contig); + // if contig is specified and we are past that contig, we are done: + if ( location != null && GenomeLoc.compareContigs(cur_contig, location.getContig()) == 1 ) break; + if ( location == null || GenomeLoc.compareContigs(cur_contig, location.getContig()) == 0 ) { + contig_seq = reference.get(r.getReferenceIndex()); + String refstr = new String(contig_seq.getBases()); + col.setReferenceSequence(refstr); + if (!CONTROL_RUN) pileBuilder.setReferenceSequence(refstr); + System.out.println("loaded contig "+cur_contig+" (index="+r.getReferenceIndex()+"); length="+contig_seq.getBases().length+" tst="+contig_seq.toString()); + } + } + + // if contig is specified and we did not reach it yet, skip the records until we reach that contig: + if ( location != null && GenomeLoc.compareContigs(cur_contig, location.getContig()) == -1 ) continue; + + + if ( location != null && r.getAlignmentEnd() < location.getStart() ) continue; + + // if stop position is specified and we are past that, stop reading: + if ( location != null && r.getAlignmentStart() > location.getStop() ) break; + + // if ( cur_contig.equals("chrM") || GenomeLoc.compareContigs(cur_contig,"chrY") > 0 ) continue; // skip chrM and unplaced contigs for now + + // we currently do not know how to deal with cigars containing elements other than M,I,D, so + // let's just skip the reads that contain those other elements (clipped reads?) + Cigar c = r.getCigar(); + boolean cigar_acceptable = true; + for ( int z = 0 ; z < c.numCigarElements() ; z++ ) { + CigarElement ce = c.getCigarElement(z); + switch ( ce.getOperator() ) { + case M: + case I: + case D: break; + default: + cigar_acceptable = false; + } + } + if ( ! cigar_acceptable ) { + discarded_cigar_count++; + continue; + } + + if ( r.getReadLength() > MAX_READ_LENGTH ) { + discarded_long_read_count++; + continue; + } + + int err = -1; +/* + System.out.println("MM: "+numMismatches(r)); + System.out.println("direct: "+numMismatchesDirect(r,contig_seq)); + System.out.print(" "); + for ( int i = r.getAlignmentStart() - 1 ; i < r.getAlignmentEnd() ; i++ ) System.out.print((char)contig_seq.getBases()[i]); + System.out.println(); + System.out.println((r.getReadNegativeStrandFlag()?"<-":"->")+r.getReadString()); + System.out.println("cigar: "+r.getCigarString()); + System.out.println(); + if (counter++ == 20 ) break; + continue; +*/ + + if ( ERR_MODE.equals("MM")) err = numMismatches(r,contig_seq); + else if ( ERR_MODE.equals("MC") ) err = AlignmentUtils.numMismatches(r,contig_seq); + else if ( ERR_MODE.equals("ERR")) err = numErrors(r,contig_seq); + else if ( ERR_MODE.equals("MG")) err = numMismatchesGaps(r,contig_seq); + if ( err > MAX_ERRS.intValue() ) continue; + // counter++; + // if ( counter % 1000000 == 0 ) System.out.println(counter+" records; "+col.memStatsString()); + col.receive(r); + + } + + if ( ! CONTROL_RUN ) { + pileBuilder.printStats(); + pileBuilder.close(); + } + System.out.println("done."); + System.out.println("Discarded reads with non-M,I,D cigar elements: "+ discarded_cigar_count); + System.out.println("Discarded long reads (above "+MAX_READ_LENGTH+" bp): "+ discarded_long_read_count); + System.out.println(); + col.printLengthHistograms(); + samReader.close(); + ptWriter.close(); + return 0; + } + + /** This method is a HACK: it is designed to work around the current bug in NM tags created at CRD + * + * @param r SAM record that must specify an alignment + * @return number of errors (number of mismatches plus total length of all insertions/deletions + * @throws RuntimeException + */ + private static int numErrors(SAMRecord r, ReferenceSequence refseq) throws RuntimeException { + + // NM currently stores the total number of mismatches in all blocks + 1 + int errs = numMismatches(r,refseq); + + // now we have to add the total length of all indels: + Cigar c = r.getCigar(); + for ( int i = 0 ; i < c.numCigarElements() ; i++ ) { + CigarElement ce = c.getCigarElement(i); + switch( ce.getOperator()) { + case M : break; // we already have correct number of mismatches + case I : + case D : + errs += ce.getLength(); + break; + default: throw new RuntimeException("Unrecognized cigar element"); + } + } + return errs; + } + + /** This method is a HACK: it is designed to work around the current bug in NM tags created at CRD + * + * @param r SAM record that must specify an alignment + * @return number of errors (number of mismatches plus total number of all insertions/deletions (each insertion or + * deletion will be counted as a single error regardless of the length) + * @throws RuntimeException + */ + private static int numMismatchesGaps(SAMRecord r,ReferenceSequence refseq) throws RuntimeException { + + // NM currently stores the total number of mismatches in all blocks + 1 + int errs = numMismatches(r,refseq); + + // now we have to add the total length of all indels: + Cigar c = r.getCigar(); + for ( int i = 0 ; i < c.numCigarElements() ; i++ ) { + CigarElement ce = c.getCigarElement(i); + switch( ce.getOperator()) { + case M : break; // we already have correct number of mismatches + case I : + case D : + errs++; + break; + default: throw new RuntimeException("Unrecognized cigar element"); + } + } + return errs; + } + + /** This method is a HACK: it is designed to work around the current bug in NM tags created at CRD */ + private static int numMismatches(SAMRecord r, ReferenceSequence refseq) throws RuntimeException { + + // NM currently stores the total number of mismatches in all blocks + 1 + Integer i = (Integer)r.getAttribute("NM"); + if ( i == null ) return AlignmentUtils.numMismatches(r,refseq); + return ((Integer)r.getAttribute("NM")).intValue() - 1; + + } + + /** Trivial utility method that goes some distance trying to ensure that the input file is there; + * the only purpose is reducing clutter in main(). Receives a default + * input file argument, does a few checks (e.g. that it is non-null and exists), if they fail tries + * to fire up a file chooser dialog using start_folder as initial directory, etc. + * @param default_arg some "default" input file; if it is non-null and exists, nothing else will be done, + * and the same default_arg objetc will be returned; otherwise the method will try to ask for a "better" input. + * @param start_folder should file open dialog be fired up, it will initially display this directory. + * @return File object that is not null and does exist (there is no check that it is a valid SAM/BAM file though). + */ + private File getInputFile(File default_arg, String start_folder) { + File f = default_arg; + if ( f==null || ! f.exists() ) { + JFileChooser fc = new JFileChooser(start_folder); + FileNameExtensionFilter ff = new FileNameExtensionFilter("SAM and BAM files","sam","bam"); + fc.setFileFilter(ff); + fc.setFileSelectionMode(JFileChooser.FILES_ONLY); + + int ret = fc.showOpenDialog(null); + f = fc.getSelectedFile(); + if ( ret != JFileChooser.APPROVE_OPTION ) { + System.out.println("No input file specified. Exiting..."); + System.exit(1); + } + } + + if ( f == null || ! f.exists() ) { + System.out.println("SAM or BAM input file must be specified. Exiting..."); + System.exit(1); + } + + return f; + } + + /** Auxiliary method to remove some clutter from main(); gets called only once and tries to get + * contig ordering from the header provided by opened SAM reader; if no header info is available + * falls back to default ordering; whichever ordering is used, it is set for GenomeLoc class. + * @param r sam reader to get header from + */ + private void setContigOrdering(SAMFileReader r) { + SAMFileHeader h = r.getFileHeader(); + if ( h == null ) { + System.out.println("No header found in SAM file, falling back to default contig ordering"); + setDefaultContigOrdering(); + return; + } + GenomeLoc.setupRefContigOrdering(h.getSequenceDictionary()); + } + + private void setDefaultContigOrdering() { + Map rco = new HashMap(); + rco.put("chrM",0); + for ( int i = 1 ; i <= 22 ; i++ ) rco.put(Integer.toString(i),i);//rco.put("chr"+i,i); + rco.put("chrX",23); + rco.put("chrY",24); + } +}