int DataStatisticsTool::Execute()
{
// iterate over reads in BAM file(s)
BamAlignment alignObj;
while(bamReader.GetNextAlignment(alignObj))
{
if (alignObj.IsDuplicate()) continue;
if (alignObj.IsFailedQC()) continue;
if (!alignObj.IsMapped()) continue;
if (!alignObj.IsPrimaryAlignment()) continue;
if (alignObj.IsPaired() && !alignObj.IsProperPair()) continue;
if (alignObj.IsPaired() && !alignObj.IsMateMapped()) continue;
if (!alignObj.HasTag("MD")) continue;
// // debug
// GenericBamAlignmentTools::printBamAlignmentCigar(alignObj);
// GenericBamAlignmentTools::printBamAlignmentMD(alignObj);
// shift InDel
GenericBamAlignmentTools::leftShiftInDel(alignObj);
// // debug
// GenericBamAlignmentTools::printBamAlignmentCigar(alignObj);
// GenericBamAlignmentTools::printBamAlignmentMD(alignObj);
// get the alignment sequences
string alignRead;
string alignGenome;
GenericBamAlignmentTools::getAlignmentSequences(alignObj, alignRead, alignGenome);
// update the statistics
statistics.update(alignRead, alignGenome);
}
// print to screen
cout << statistics << endl;
// statistics.printMatchMismatch();
// close BAM reader
bamReader.Close();
// close Fasta
genomeFasta.Close();
return 1;
}
// use current input alignment to update BAM file alignment stats
void StatsTool::StatsToolPrivate::ProcessAlignment(const BamAlignment& al) {
// increment total alignment counter
++numReads;
// check the paired-independent flags
if ( al.IsDuplicate() ) ++numDuplicates;
if ( al.IsFailedQC() ) ++numFailedQC;
if ( al.IsMapped() ) ++numMapped;
// check forward/reverse strand
if ( al.IsReverseStrand() )
++numReverseStrand;
else
++numForwardStrand;
// if alignment is paired-end
if ( al.IsPaired() ) {
// increment PE counter
++numPaired;
// increment first mate/second mate counters
if ( al.IsFirstMate() ) ++numFirstMate;
if ( al.IsSecondMate() ) ++numSecondMate;
// if alignment is mapped, check mate status
if ( al.IsMapped() ) {
// if mate mapped
if ( al.IsMateMapped() )
++numBothMatesMapped;
// else singleton
else
++numSingletons;
}
// check for explicit proper pair flag
if ( al.IsProperPair() ) ++numProperPair;
// store insert size for first mate
if ( settings->IsShowingInsertSizeSummary && al.IsFirstMate() && (al.InsertSize != 0) ) {
int insertSize = abs(al.InsertSize);
insertSizes.push_back( insertSize );
}
}
}
bool check(const PropertyFilter& filter, const BamAlignment& al) {
bool keepAlignment = true;
const PropertyMap& properties = filter.Properties;
PropertyMap::const_iterator propertyIter = properties.begin();
PropertyMap::const_iterator propertyEnd = properties.end();
for ( ; propertyIter != propertyEnd; ++propertyIter ) {
// check alignment data field depending on propertyName
const string& propertyName = (*propertyIter).first;
const PropertyFilterValue& valueFilter = (*propertyIter).second;
if ( propertyName == ALIGNMENTFLAG_PROPERTY ) keepAlignment &= valueFilter.check(al.AlignmentFlag);
else if ( propertyName == CIGAR_PROPERTY ) {
stringstream cigarSs;
const vector<CigarOp>& cigarData = al.CigarData;
if ( !cigarData.empty() ) {
vector<CigarOp>::const_iterator cigarBegin = cigarData.begin();
vector<CigarOp>::const_iterator cigarIter = cigarBegin;
vector<CigarOp>::const_iterator cigarEnd = cigarData.end();
for ( ; cigarIter != cigarEnd; ++cigarIter ) {
const CigarOp& op = (*cigarIter);
cigarSs << op.Length << op.Type;
}
keepAlignment &= valueFilter.check(cigarSs.str());
}
}
else if ( propertyName == INSERTSIZE_PROPERTY ) keepAlignment &= valueFilter.check(al.InsertSize);
else if ( propertyName == ISDUPLICATE_PROPERTY ) keepAlignment &= valueFilter.check(al.IsDuplicate());
else if ( propertyName == ISFAILEDQC_PROPERTY ) keepAlignment &= valueFilter.check(al.IsFailedQC());
else if ( propertyName == ISFIRSTMATE_PROPERTY ) keepAlignment &= valueFilter.check(al.IsFirstMate());
else if ( propertyName == ISMAPPED_PROPERTY ) keepAlignment &= valueFilter.check(al.IsMapped());
else if ( propertyName == ISMATEMAPPED_PROPERTY ) keepAlignment &= valueFilter.check(al.IsMateMapped());
else if ( propertyName == ISMATEREVERSESTRAND_PROPERTY ) keepAlignment &= valueFilter.check(al.IsMateReverseStrand());
else if ( propertyName == ISPAIRED_PROPERTY ) keepAlignment &= valueFilter.check(al.IsPaired());
else if ( propertyName == ISPRIMARYALIGNMENT_PROPERTY ) keepAlignment &= valueFilter.check(al.IsPrimaryAlignment());
else if ( propertyName == ISPROPERPAIR_PROPERTY ) keepAlignment &= valueFilter.check(al.IsProperPair());
else if ( propertyName == ISREVERSESTRAND_PROPERTY ) keepAlignment &= valueFilter.check(al.IsReverseStrand());
else if ( propertyName == ISSECONDMATE_PROPERTY ) keepAlignment &= valueFilter.check(al.IsSecondMate());
else if ( propertyName == ISSINGLETON_PROPERTY ) {
const bool isSingleton = al.IsPaired() && al.IsMapped() && !al.IsMateMapped();
keepAlignment &= valueFilter.check(isSingleton);
}
else if ( propertyName == MAPQUALITY_PROPERTY ) keepAlignment &= valueFilter.check(al.MapQuality);
else if ( propertyName == MATEPOSITION_PROPERTY ) keepAlignment &= ( al.IsPaired() && al.IsMateMapped() && valueFilter.check(al.MateRefID) );
else if ( propertyName == MATEREFERENCE_PROPERTY ) {
if ( !al.IsPaired() || !al.IsMateMapped() ) return false;
BAMTOOLS_ASSERT_MESSAGE( (al.MateRefID>=0 && (al.MateRefID<(int)filterToolReferences.size())), "Invalid MateRefID");
const string& refName = filterToolReferences.at(al.MateRefID).RefName;
keepAlignment &= valueFilter.check(refName);
}
else if ( propertyName == NAME_PROPERTY ) keepAlignment &= valueFilter.check(al.Name);
else if ( propertyName == POSITION_PROPERTY ) keepAlignment &= valueFilter.check(al.Position);
else if ( propertyName == QUERYBASES_PROPERTY ) keepAlignment &= valueFilter.check(al.QueryBases);
else if ( propertyName == REFERENCE_PROPERTY ) {
BAMTOOLS_ASSERT_MESSAGE( (al.RefID>=0 && (al.RefID<(int)filterToolReferences.size())), "Invalid RefID");
const string& refName = filterToolReferences.at(al.RefID).RefName;
keepAlignment &= valueFilter.check(refName);
}
else if ( propertyName == TAG_PROPERTY ) keepAlignment &= checkAlignmentTag(valueFilter, al);
else BAMTOOLS_ASSERT_UNREACHABLE;
// if alignment fails at ANY point, just quit and return false
if ( !keepAlignment ) return false;
}
BAMTOOLS_ASSERT_MESSAGE( keepAlignment, "Error in BamAlignmentChecker... keepAlignment should be true here");
return keepAlignment;
}
void BedGenomeCoverage::CoverageBam(string bamFile) {
ResetChromCoverage();
// open the BAM file
BamReader reader;
if (!reader.Open(bamFile)) {
cerr << "Failed to open BAM file " << bamFile << endl;
exit(1);
}
// get header & reference information
string header = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// load the BAM header references into a BEDTools "genome file"
_genome = new GenomeFile(refs);
// convert each aligned BAM entry to BED
// and compute coverage on B
BamAlignment bam;
while (reader.GetNextAlignment(bam)) {
// skip if the read is unaligned
if (bam.IsMapped() == false)
continue;
bool _isReverseStrand = bam.IsReverseStrand();
//changing second mate's strand to opposite
if( _dUTP && bam.IsPaired() && bam.IsMateMapped() && bam.IsSecondMate())
_isReverseStrand = !bam.IsReverseStrand();
// skip if we care about strands and the strand isn't what
// the user wanted
if ( (_filterByStrand == true) &&
((_requestedStrand == "-") != _isReverseStrand) )
continue;
// extract the chrom, start and end from the BAM alignment
string chrom(refs.at(bam.RefID).RefName);
CHRPOS start = bam.Position;
CHRPOS end = bam.GetEndPosition(false, false) - 1;
// are we on a new chromosome?
if ( chrom != _currChromName )
StartNewChrom(chrom);
if(_pair_chip_) {
// Skip if not a proper pair
if (bam.IsPaired() && (!bam.IsProperPair() or !bam.IsMateMapped()) )
continue;
// Skip if wrong coordinates
if( ( (bam.Position<bam.MatePosition) && bam.IsReverseStrand() ) ||
( (bam.MatePosition < bam.Position) && bam.IsMateReverseStrand() ) ) {
//chemically designed: left on positive strand, right on reverse one
continue;
}
/*if(_haveSize) {
if (bam.IsFirstMate() && bam.IsReverseStrand()) { //put fragmentSize in to the middle of pair end_fragment
int mid = bam.MatePosition+abs(bam.InsertSize)/2;
if(mid<_fragmentSize/2)
AddCoverage(0, mid+_fragmentSize/2);
else
AddCoverage(mid-_fragmentSize/2, mid+_fragmentSize/2);
}
else if (bam.IsFirstMate() && bam.IsMateReverseStrand()) { //put fragmentSize in to the middle of pair end_fragment
int mid = start+abs(bam.InsertSize)/2;
if(mid<_fragmentSize/2)
AddCoverage(0, mid+_fragmentSize/2);
else
AddCoverage(mid-_fragmentSize/2, mid+_fragmentSize/2);
}
} else */
if (bam.IsFirstMate() && bam.IsReverseStrand()) { //prolong to the mate to the left
AddCoverage(bam.MatePosition, end);
}
else if (bam.IsFirstMate() && bam.IsMateReverseStrand()) { //prolong to the mate to the right
AddCoverage(start, start + abs(bam.InsertSize) - 1);
}
} else if (_haveSize) {
if(bam.IsReverseStrand()) {
if(end<_fragmentSize) { //sometimes fragmentSize is bigger :(
AddCoverage(0, end);
} else {
AddCoverage(end + 1 - _fragmentSize, end );
}
} else {
AddCoverage(start,start+_fragmentSize - 1);
}
} else
// add coverage accordingly.
if (!_only_5p_end && !_only_3p_end) {
bedVector bedBlocks;
// we always want to split blocks when a D CIGAR op is found.
// if the user invokes -split, we want to also split on N ops.
if (_obeySplits) { // "D" true, "N" true
GetBamBlocks(bam, refs.at(bam.RefID).RefName, bedBlocks, true, true);
}
else { // "D" true, "N" false
GetBamBlocks(bam, refs.at(bam.RefID).RefName, bedBlocks, true, false);
}
AddBlockedCoverage(bedBlocks);
}
else if (_only_5p_end) {
CHRPOS pos = ( !bam.IsReverseStrand() ) ? start : end;
AddCoverage(pos,pos);
}
else if (_only_3p_end) {
CHRPOS pos = ( bam.IsReverseStrand() ) ? start : end;
AddCoverage(pos,pos);
}
}
// close the BAM
reader.Close();
// process the results of the last chromosome.
ReportChromCoverage(_currChromCoverage, _currChromSize,
_currChromName, _currChromDepthHist);
// report all empty chromsomes
PrintEmptyChromosomes();
// report the overall coverage if asked.
PrintFinalCoverage();
}
