// close the BAM files
void BamMultiReader::Close(void) {
// close all BAM readers and clean up pointers
vector<pair<BamReader*, BamAlignment*> >::iterator readerIter = readers.begin();
vector<pair<BamReader*, BamAlignment*> >::iterator readerEnd = readers.end();
for ( ; readerIter != readerEnd; ++readerIter) {
BamReader* reader = (*readerIter).first;
BamAlignment* alignment = (*readerIter).second;
// close the reader
if ( reader) reader->Close();
// delete reader pointer
delete reader;
reader = 0;
// delete alignment pointer
delete alignment;
alignment = 0;
}
// clear out the container
readers.clear();
}
void BedIntersectPE::IntersectBamPE(string bamFile) {
// load the "B" bed file into a map so
// that we can easily compare "A" to it for overlaps
_bedB->loadBedFileIntoMap();
// open the BAM file
BamReader reader;
BamWriter writer;
reader.Open(bamFile);
// get header & reference information
string bamHeader = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// open a BAM output to stdout if we are writing BAM
if (_bamOutput == true) {
// set compression mode
BamWriter::CompressionMode compressionMode = BamWriter::Compressed;
if ( _isUncompressedBam ) compressionMode = BamWriter::Uncompressed;
writer.SetCompressionMode(compressionMode);
// open our BAM writer
writer.Open("stdout", bamHeader, refs);
}
// track the previous and current sequence
// names so that we can identify blocks of
// alignments for a given read ID.
string prevName, currName;
prevName = currName = "";
vector<BamAlignment> alignments; // vector of BAM alignments for a given ID in a BAM file.
alignments.reserve(100);
_bedA->bedType = 10; // it's a full BEDPE given it's BAM
// rip through the BAM file and convert each mapped entry to BEDPE
BamAlignment bam1, bam2;
while (reader.GetNextAlignment(bam1)) {
// the alignment must be paired
if (bam1.IsPaired() == true) {
// grab the second alignment for the pair.
reader.GetNextAlignment(bam2);
// require that the alignments are from the same query
if (bam1.Name == bam2.Name) {
ProcessBamBlock(bam1, bam2, refs, writer);
}
else {
cerr << "*****ERROR: -bedpe requires BAM to be sorted or grouped by query name. " << endl;
exit(1);
}
}
}
// close up
reader.Close();
if (_bamOutput == true) {
writer.Close();
}
}
/*
Description:
Load all the bam into memory at one time if no parameters set, otherwise load the needed part of the bam.
Save the parsed info into vector.
*/
bool BamParse::parseAlignment(int chrom1, int chrom1_begin, int chrom2, int chrom2_end)
{
BamReader reader;
if ( !reader.Open(filename) ) {
cerr << "Bamtools ERROR: could not open input BAM file: " << filename << endl;
return false;
}
//check whether need to set a region.
if(chrom1>-1 && chrom1_begin>-1 && chrom2>-1 && chrom2_end>-1)
{
this->loadIndex(reader);
BamRegion br(chrom1,chrom1_begin,chrom2,chrom2_end);
bool is_set=reader.SetRegion(br);
if(is_set==false)
{
return false;//cannot set the region.
}
}
//process input data
BamAlignment al;
while ( reader.GetNextAlignment(al) )
{
if(al.Position<0) continue;
BamAlignmentRecord* bar=new BamAlignmentRecord();
setAlignmentRecord(al,bar);
bam_aln_records.push_back(bar);
}
reader.Close();
return true;
}
int main (int argc, char *argv[]) {
string bamfiletopen = string(argv[1]);
BamReader reader;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
BamAlignment al;
while ( reader.GetNextAlignment(al) ) {
string reconstructedReference = reconstructRef(&al);
cout<<al.QueryBases<<endl;
cout<<reconstructedReference<<endl;
pair< string, vector<int> > reconP = reconstructRefWithPos(&al);
for(unsigned int i=0;i<reconP.first.size();i++){
cout<<reconP.first[i]<<"\t"<<reconP.second[i]<<endl;
}
}
reader.Close();
return 0;
}
bool RevertTool::RevertToolPrivate::Run(void) {
// opens the BAM file without checking for indexes
BamReader reader;
if ( !reader.Open(m_settings->InputFilename) ) {
cerr << "Could not open input BAM file... quitting." << endl;
return false;
}
// get BAM file metadata
const string& headerText = reader.GetHeaderText();
const RefVector& references = reader.GetReferenceData();
// open writer
BamWriter writer;
bool writeUncompressed = ( m_settings->OutputFilename == Options::StandardOut() && !m_settings->IsForceCompression );
if ( !writer.Open(m_settings->OutputFilename, headerText, references, writeUncompressed) ) {
cerr << "Could not open " << m_settings->OutputFilename << " for writing." << endl;
return false;
}
// plow through file, reverting alignments
BamAlignment al;
while ( reader.GetNextAlignment(al) ) {
RevertAlignment(al);
writer.SaveAlignment(al);
}
// clean and exit
reader.Close();
writer.Close();
return true;
}
/**
* Main work method. Reads the BAM file once and collects sorted information about
* the 5' ends of both ends of each read (or just one end in the case of pairs).
* Then makes a pass through those determining duplicates before re-reading the
* input file and writing it out with duplication flags set correctly.
*/
int MarkDuplicates::runInternal() {
ogeNameThread("am_MarkDuplicates");
if(verbose)
cerr << "Reading input file and constructing read end information." << endl;
buildSortedReadEndLists();
generateDuplicateIndexes();
if(verbose)
cerr << "Marking " << numDuplicateIndices << " records as duplicates." << endl;
BamReader in;
in.Open(getBufferFileName());
// Now copy over the file while marking all the necessary indexes as duplicates
long recordInFileIndex = 0;
long written = 0;
while (true) {
BamAlignment * prec = in.GetNextAlignment();
if(!prec)
break;
if (prec->IsPrimaryAlignment()) {
if (duplicateIndexes.count(recordInFileIndex) == 1)
prec->SetIsDuplicate(true);
else
prec->SetIsDuplicate(false);
}
recordInFileIndex++;
if (removeDuplicates && prec->IsDuplicate()) {
// do nothing
}
else {
putOutputAlignment(prec);
if (verbose && read_count && ++written % 100000 == 0) {
cerr << "\rWritten " << written << " records (" << written * 100 / read_count <<"%)." << std::flush;
}
}
}
if (verbose && read_count)
cerr << "\rWritten " << written << " records (" << written * 100 / read_count <<"%)." << endl;
in.Close();
remove(getBufferFileName().c_str());
return 0;
}
void BedCoverage::CollectCoverageBam(string bamFile) {
// load the "B" bed file into a map so
// that we can easily compare "A" to it for overlaps
_bedB->loadBedCovFileIntoMap();
// open the BAM file
BamReader reader;
reader.Open(bamFile);
// get header & reference information
string header = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// convert each aligned BAM entry to BED
// and compute coverage on B
BamAlignment bam;
while (reader.GetNextAlignment(bam)) {
if (bam.IsMapped()) {
// treat the BAM alignment as a single "block"
if (_obeySplits == false) {
// construct a new BED entry from the current BAM alignment.
BED a;
a.chrom = refs.at(bam.RefID).RefName;
a.start = bam.Position;
a.end = bam.GetEndPosition(false, false);
a.strand = "+";
if (bam.IsReverseStrand()) a.strand = "-";
_bedB->countHits(a, _sameStrand, _diffStrand, _countsOnly);
}
// split the BAM alignment into discrete blocks and
// look for overlaps only within each block.
else {
// vec to store the discrete BED "blocks" from a
bedVector bedBlocks;
// since we are counting coverage, we do want to split blocks when a
// deletion (D) CIGAR op is encountered (hence the true for the last parm)
GetBamBlocks(bam, refs.at(bam.RefID).RefName, bedBlocks, false, true);
// use countSplitHits to avoid over-counting each split chunk
// as distinct read coverage.
_bedB->countSplitHits(bedBlocks, _sameStrand, _diffStrand, _countsOnly);
}
}
}
// report the coverage (summary or histogram) for BED B.
if (_countsOnly == true)
ReportCounts();
else
ReportCoverage();
// close the BAM file
reader.Close();
}
bool CoverageTool::CoverageToolPrivate::Run(void) {
// if output filename given
ofstream outFile;
if ( m_settings->HasOutputFile ) {
// open output file stream
outFile.open(m_settings->OutputFilename.c_str());
if ( !outFile ) {
cerr << "bamtools coverage ERROR: could not open " << m_settings->OutputFilename
<< " for output" << endl;
return false;
}
// set m_out to file's streambuf
m_out.rdbuf(outFile.rdbuf());
}
//open our BAM reader
BamReader reader;
if ( !reader.Open(m_settings->InputBamFilename) ) {
cerr << "bamtools coverage ERROR: could not open input BAM file: " << m_settings->InputBamFilename << endl;
return false;
}
// retrieve references
m_references = reader.GetReferenceData();
// set up our output 'visitor'
CoverageVisitor* cv = new CoverageVisitor(m_references, &m_out);
// set up pileup engine with 'visitor'
PileupEngine pileup;
pileup.AddVisitor(cv);
// process input data
BamAlignment al;
while ( reader.GetNextAlignment(al) )
pileup.AddAlignment(al);
// clean up
reader.Close();
if ( m_settings->HasOutputFile )
outFile.close();
delete cv;
cv = 0;
// return success
return true;
}
int main (int argc, char *argv[]) {
if( (argc== 1) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cout<<"Usage:setAsUnpaired [in bam] [outbam]"<<endl<<"this program takes flags all paired sequences as singles"<<endl;
return 1;
}
string bamfiletopen = string(argv[1]);
string bamFileOUT = string(argv[2]);
BamReader reader;
BamWriter writer;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
const SamHeader header = reader.GetHeader();
const RefVector references = reader.GetReferenceData();
if ( !writer.Open(bamFileOUT,header,references) ) {
cerr << "Could not open output BAM file "<<bamFileOUT << endl;
return 1;
}
BamAlignment al;
while ( reader.GetNextAlignment(al) ) {
if(al.IsMapped()){
cerr << "Cannot yet handle mapped reads " << endl;
return 1;
}
al.SetIsPaired (false);
writer.SaveAlignment(al);
} //while al
reader.Close();
writer.Close();
return 0;
}
int IndexTool::Run(int argc, char* argv[]) {
// parse command line arguments
Options::Parse(argc, argv, 1);
// open our BAM reader
BamReader reader;
reader.Open(m_settings->InputBamFilename);
// create index for BAM file
bool useDefaultIndex = !m_settings->IsUsingBamtoolsIndex;
reader.CreateIndex(useDefaultIndex);
// clean & exit
reader.Close();
return 0;
}
int main (int argc, char *argv[]) {
if( (argc== 1) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cout<<"Usage:editDist [in bam]"<<endl<<"this program returns the NM field of all aligned reads"<<endl;
return 1;
}
string bamfiletopen = string(argv[1]);
// cout<<bamfiletopen<<endl;
BamReader reader;
// cout<<"ok"<<endl;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
BamAlignment al;
// cout<<"ok"<<endl;
while ( reader.GetNextAlignment(al) ) {
// cout<<al.Name<<endl;
if(!al.IsMapped())
continue;
if(al.HasTag("NM") ){
int editDist;
if(al.GetTag("NM",editDist) ){
cout<<editDist<<endl;
}else{
cerr<<"Cannot retrieve NM field for "<<al.Name<<endl;
return 1;
}
}else{
cerr<<"Warning: read "<<al.Name<<" is aligned but has no NM field"<<endl;
}
} //while al
reader.Close();
return 0;
}
int main (int argc, char** argv)
{
// Print Commandline
string ss(argv[0]); // convert Char to String
string commandline = "##Print Command line " + ss;
int c;
FastaReference* reference = NULL;
int minbaseQ = 10; //default
int windowlen = 40; //by default
string regionstr;
string RegionFile;
string bamfile;
bool STdin = false;
bool has_region = false;
bool has_regionFile = false;
bool has_bamfile = false;
bool has_ref = false;
int ploidy = 2;
bool SetLowComplexityRegionSWGapExt = false;
bool SetLowComplexityRegion = false;
if (argc < 2)
{
printSummary(argv);
exit(1);
}
while (true)
{
static struct option long_options[] =
{
{"help", no_argument, 0, 'h'},
{"ploidy", required_argument, 0, 'p'},
{"window-size", required_argument, 0, 'w'},
{"reference", required_argument, 0, 'f'},
{"min-base-quality", required_argument, 0,'q'},
{"Region", required_argument, 0, 'R'},
{"STdin", no_argument, 0, 's'},
{"bam", required_argument, 0, 'b'},
{"Repeat-Extgap", no_argument, 0, 'E'},
{"LowCompex", no_argument, 0, 'l'},
{0, 0, 0, 0}
};
int option_index = 0;
c = getopt_long (argc, argv, "hslEf:q:w:s:r:R:p:b:", long_options, &option_index);
/* Detect the end of the options. */
if (c == -1)
break;
switch (c)
{
case 'f':
reference = new FastaReference(optarg); // will exit on open failure
commandline = commandline + " -f " + optarg;
has_ref = true;
break;
case 'b':
has_bamfile = true;
bamfile = optarg;
commandline = commandline + " -b " + optarg;
break;
case 'r':
regionstr = optarg;
has_region = true;
commandline = commandline + " -r " + optarg;
break;
case 'R':
RegionFile = optarg;
has_regionFile = true;
commandline = commandline + " -R " + optarg;
break;
case 's':
STdin = true;
commandline = commandline + " -s ";
break;
case 'q':
minbaseQ = atoi(optarg);
commandline = commandline + " -q " + optarg;
break;
case 'w':
windowlen = atoi(optarg);
commandline = commandline + " -w " + optarg;
break;
case 'p':
ploidy = atoi(optarg);
commandline = commandline + " -p " + optarg;
break;
case 'E':
SetLowComplexityRegionSWGapExt = true;
commandline = commandline + " -E ";
break;
case 'l':
SetLowComplexityRegion = true;
commandline = commandline + " -l ";
break;
case 'h':
printSummary(argv);
commandline = commandline + " -h ";
exit(0);
break;
case '?':
printSummary(argv);
exit(1);
break;
default:
abort();
break;
}
}
//// Open Error log files
ofstream cerrlog("bonsaiReport.txt");
streambuf *cerrsave = std::cerr.rdbuf();
// Redirect stream buffers
if (cerrlog.is_open())
cerr.rdbuf(cerrlog.rdbuf());
cerr << commandline << endl;
//Check for Reference Fasta sequence
if (!has_ref)
{
cerr << "no FASTA reference provided, cannot realign" << endl;
exit(1);
}
////Check for reader
BamReader reader;
if (STdin == true)
{
if (!reader.Open("stdin"))
{
cerr << "could not open stdin bam for reading" << endl;
cerr << reader.GetErrorString() << endl;
reader.Close();
printSummary(argv);
}
}
else
{
if (has_bamfile == true)
{
if (!reader.Open(bamfile))
{
cerr << "ERROR: could not open bam files from stdin ... Aborting" << endl;
cerr << reader.GetErrorString() << endl;
reader.Close();
printSummary(argv);
}
if ( !reader.LocateIndex() )
reader.CreateIndex();
}
else
{
cerr << "--bam flag is set but no bamfile is provided... Aborting" << endl;
reader.Close();
printSummary(argv);
}
}
//// Check Region Tags
if ( (has_regionFile == true) && (has_region == true) )
{
cerr << "ERROR: You provide both region and has provide a Set Region List... Aborting" << endl;
exit(1);
}
//// store the names of all the reference sequences in the BAM file
vector<RefData> referencedata = reader.GetReferenceData();
//// Store Region LIST
vector<BamRegion> regionlist;
if (has_region == true)
{
BamRegion region;
ParseRegionString(regionstr, reader, region);
regionlist.push_back(region);
}
else if (has_regionFile == true)
{
ifstream RG(RegionFile.c_str(), ios_base::in);
string line;
while(getline(RG,line))
{
BamRegion region;
ParseRegionString(line, reader, region);
regionlist.push_back(region);
}
RG.close();
}
else if ( (has_regionFile == false) && (has_region == false) )
{
for (int i= 0; i < (int)referencedata.size(); i++)
{
string regionstr = referencedata.at(i).RefName;
BamRegion region;
ParseRegionString(regionstr, reader, region);
if (!reader.SetRegion(region)) // Bam region will get [0,101) = 0 to 100 => [closed, half-opened)
{
cerr << "ERROR: set region " << regionstr << " failed. Check that REGION describes a valid range... Aborting" << endl;
reader.Close();
exit(1);
}
else
regionlist.push_back(region);
}
}
////
BamWriter writer;
if (!writer.Open("stdout", reader.GetHeaderText(), reader.GetReferenceData()))
{
cerr << "could not open stdout for writing" << endl;
exit(1);
}
//// Smallest start position and Largest end position for Req Seq
vector<RefData>::iterator refdataIter = referencedata.begin();
vector<BamRegion>::iterator regionListIter = regionlist.begin();
// CLASS
RealignFunctionsClass RealignFunction;
map<int, string> RefIDRedName;
vector<SalRealignInfo> AlGroups;
multimap<int, BamAlignment> SortRealignedAlignmentsMultimap;
int refid = 0;
BamAlignment alignment;
bool IsNextAlignment = reader.GetNextAlignment(alignment);
//cerr << " " << alignment.Name << " Chr " << alignment.RefID << " Startpos: " << alignment.Position << " Endpos: " << alignment.GetEndPosition() << " Length: " << alignment.Length << endl;
int windowrealigned = 0;
int TotalWindowDetected = 0;
int TotalReadsAligned = 0;
int TotalWindow = 0;
int TotalReads = 0;
while (refdataIter != referencedata.end() )
{
string refname = refdataIter->RefName;
RefIDRedName[refid] = refname;
int reflength = refdataIter->RefLength;
int winstartpos, winendpos;
int AllowableBasesInWindow = 1;
bool nextChrName = false;
cerr << "##HeaderINFO: RefID = " << refdataIter->RefName << "\t" << "RefLen = " << reflength << endl;
while (nextChrName == false )
{
vector<int> minmaxRefSeqPos;
bool IsPassDetectorNoRealignment = false;
minmaxRefSeqPos.push_back(-1);
minmaxRefSeqPos.push_back(0);
//cerr << " region: " << (*regionListIter).LeftRefID << " : " << (*regionListIter).LeftPosition << " .. " << (*regionListIter).RightPosition << endl;
if ((refid == (int)referencedata.size() - 1) && ((*regionListIter).LeftRefID == refid) && ((has_region==true) || (has_regionFile==true)) )
{
////
if ( (has_region == true) || (has_regionFile == true) )
{
winstartpos = (*regionListIter).LeftPosition;
winendpos = winstartpos + windowlen - 1;
reflength = (*regionListIter).RightPosition;
if (reflength < winendpos)
reflength = winendpos;
// Get Next Alignment First
if ( (refid == alignment.RefID) && (winstartpos == (*regionListIter).LeftPosition) && (IsNextAlignment == false) )
IsNextAlignment = reader.GetNextAlignment(alignment);
}
else if (has_region == false)
{
winstartpos = 0;
winendpos = winstartpos + windowlen - 1;
// Get Next Alignment First
if ( (refid == alignment.RefID) && (winstartpos == 0) && (IsNextAlignment == false) )
IsNextAlignment = reader.GetNextAlignment(alignment);
}
//cerr << " winstart: " << winstartpos << " ; winend: " << winendpos;
//cerr << " " << alignment.Name << " Chr " << alignment.RefID << " Startpos: " << alignment.Position << " Endpos: " << alignment.GetEndPosition() << " Length: " << alignment.Length << endl;
////
while ((winstartpos < reflength))
{
//// Check window end position
if (winendpos > reflength)
winendpos = reflength;
// Reinitialized
unsigned int NewReadMappedcount = 0;
//// Save and Erase alignments that are outside of window (Deque?)
if (!AlGroups.empty())
{
minmaxRefSeqPos.at(0) = -1;
minmaxRefSeqPos.at(1) = 0;
//cerr << "#Start: Keep alignments with start position exceed the right end of the window/Region " << endl;
vector<SalRealignInfo>::iterator Iter = AlGroups.begin();
while (Iter != AlGroups.end())
{
// Erase alignment s
if ((*Iter).al.GetEndPosition() < winstartpos)
{
//cerr << " ToWrite: " << (*Iter).second.size() << " ; " << (*Iter).al.Name << " ; " << (*Iter).al.Position << " < " << winstartpos << " : " << (*Iter).al.GetEndPosition() << endl;
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > ((*Iter).al.Position, (*Iter).al));
AlGroups.erase(Iter);
//cerr << " ToWrite: DONE " << endl;
}
else
{
string referenceSequence = reference->getSubSequence(RefIDRedName[(*Iter).al.RefID], (*Iter).al.Position, 2*(*Iter).al.Length);
if ((*Iter).HasRealign == true )
{
(*Iter).currentReadPosition = 0;
(*Iter).currentGenomeSeqPosition = 0;
(*Iter).currentAlPosition = (*Iter).al.Position;
(*Iter).cigarindex = 0;
}
(*Iter).CigarSoftclippingLength = 0;
SalRealignInfo talr = (*Iter);
//cerr << " ToKEEP: " << (*Iter).al.Name << " ; " << (*Iter).al.Position << " < " << winstartpos << " : " << (*Iter).al.GetEndPosition() << endl;
RealignFunction.ParseAlignmentsAndExtractVariantsByBaseQualv(AlGroups, talr, Iter, (*Iter).al, referenceSequence, minmaxRefSeqPos, winstartpos, winendpos, (float) minbaseQ, false);
++Iter; //Increment iterator
}
}
}
// Write Sorted Alignments that are outside of window
//cerr << "SortRealignedAlignmentsMultimap: " << SortRealignedAlignmentsMultimap.size() << " minmaxRefSeqPos.at(0)= " << minmaxRefSeqPos.at(0) << endl;
if (!SortRealignedAlignmentsMultimap.empty()) // && (winWrite < winstartpos ) )
{
//cerr << "#Start: Write alignments and delete alignments with start position exceed the right end of the window/Region " << endl;
multimap<int, BamAlignment>::iterator sraIter = SortRealignedAlignmentsMultimap.begin();
while (sraIter != SortRealignedAlignmentsMultimap.end())
{
//cerr << " (*sraIter).first= " << (*sraIter).first << " minmaxRefSeqPos.at(0)= " << minmaxRefSeqPos.at(0) << " winstartpos - ((windowlen - 1)*0.9)= " << winstartpos - ((windowlen - 1)*0.9) << endl;
if (((float) (*sraIter).first < floor((float) (winstartpos - ((windowlen - 1)*0.9)))) && ((minmaxRefSeqPos.at(0) > 0) && ((*sraIter).first < minmaxRefSeqPos.at(0)))) {
//writer.SaveAlignment((*sraIter).second); // Why sometimes, it doesn't work ?????
if (!writer.SaveAlignment((*sraIter).second))
cerr << writer.GetErrorString() << endl;
SortRealignedAlignmentsMultimap.erase(sraIter++);
} else {
++sraIter;
}
}
//cerr << "#Done: Write alignments and delete alignments with start position exceed the right end of the window/Region " << endl;
}
//cerr << " winstart: " << winstartpos << " ; winend: " << winendpos;
//cerr << " " << alignment.Name << " Chr " << alignment.RefID << " Startpos: " << alignment.Position << " Endpos: " << alignment.GetEndPosition() << " Length: " << alignment.Length << endl;
//cerr << ": " << alignment.RefID << " :" << RefIDRedName[alignment.RefID] << " : " << RefIDRedName[alignment.RefID] << endl;
//cerr << "Start: Gather aligmenets that lie (fully or partially) within the window frame and group INDELs if there are ... " << endl;
// Gather Reads within a window frame
while ((IsNextAlignment) && (refid == alignment.RefID)) // Neeed more conditions
{
if (SetLowComplexityRegion == true)
{
string sequenceInWindow = reference->getSubSequence(RefIDRedName[alignment.RefID], winstartpos, (winendpos-winstartpos+1) );
if (IsWindowInRepeatRegion(sequenceInWindow) == true)
{
if ((IsWithinWindow(alignment, winstartpos, winendpos, AllowableBasesInWindow)) == 0)
{
TotalReads++;
if (alignment.IsMapped())
{
string referenceSequence = reference->getSubSequence(RefIDRedName[alignment.RefID], alignment.Position, 2*alignment.Length);
vector<SalRealignInfo>::iterator tIter;
SalRealignInfo alr;
alr.al = alignment;
alr.currentReadPosition = 0;
alr.currentGenomeSeqPosition = 0;
alr.currentAlPosition = alignment.Position;
alr.cigarindex = 0;
alr.HasRealign = false;
alr.CigarSoftclippingLength = 0;
string str = "ZZZZZZZZZZZZZZZZZ";
if (alignment.Name.find(str) != string::npos) {
stringstream cigar;
for (vector<CigarOp>::const_iterator cigarIter = alignment.CigarData.begin(); cigarIter != alignment.CigarData.end(); ++cigarIter)
cigar << cigarIter->Length << cigarIter->Type;
string cigarstr = cigar.str();
cerr << " TRACKING: " << alignment.RefID << " " << alignment.Name << " pos: " << alignment.Position << " cigar: " << cigarstr << endl;
}
RealignFunction.ParseAlignmentsAndExtractVariantsByBaseQualv(AlGroups, alr, tIter, alignment, referenceSequence, minmaxRefSeqPos, winstartpos, winendpos, (float) minbaseQ, true);
NewReadMappedcount++;
}
else
{
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > (alignment.Position, alignment));
cerr << "UNmapped : " << alignment.Name << endl;
}
}
else if ((IsWithinWindow(alignment, winstartpos, winendpos, AllowableBasesInWindow)) == 1)
{
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > (alignment.Position, alignment));
}
else
break;
} else {
if ((IsWithinWindow(alignment, winstartpos, winendpos, AllowableBasesInWindow)) < 2)
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > (alignment.Position, alignment));
else
break;
}
}
else // (SetLowComplexityRegion == false)
{
if ((IsWithinWindow(alignment, winstartpos, winendpos, AllowableBasesInWindow)) == 0)
{
TotalReads++;
if (alignment.IsMapped())
{
string referenceSequence = reference->getSubSequence(RefIDRedName[alignment.RefID], alignment.Position, 2 * alignment.Length);
vector<SalRealignInfo>::iterator tIter;
SalRealignInfo alr;
alr.al = alignment;
alr.currentReadPosition = 0;
alr.currentGenomeSeqPosition = 0;
alr.currentAlPosition = alignment.Position;
alr.cigarindex = 0;
alr.HasRealign = false;
alr.CigarSoftclippingLength = 0;
string str = "ZZZZZZZZZZZZZZZZZ";
if (alignment.Name.find(str) != string::npos)
{
stringstream cigar;
for (vector<CigarOp>::const_iterator cigarIter = alignment.CigarData.begin(); cigarIter != alignment.CigarData.end(); ++cigarIter)
cigar << cigarIter->Length << cigarIter->Type;
string cigarstr = cigar.str();
cerr << " TRACKING: " << alignment.RefID << " " << alignment.Name << " pos: " << alignment.Position << " cigar: " << cigarstr << endl;
}
RealignFunction.ParseAlignmentsAndExtractVariantsByBaseQualv(AlGroups, alr, tIter, alignment, referenceSequence, minmaxRefSeqPos, winstartpos, winendpos, (float) minbaseQ, true);
//cerr << " winstart: " << winstartpos << " ; winend: " << winendpos;
//cerr << " INDEL: " << alignment.RefID << " " << alignment.Name << " pos: " << alignment.Position << " Length: " << alignment.Length << " CIGARstr: " << cigarstr << endl;
NewReadMappedcount++;
}
else
{
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > (alignment.Position, alignment));
cerr << "UNmapped : " << alignment.Name << endl;
}
}
else if ((IsWithinWindow(alignment, winstartpos, winendpos, AllowableBasesInWindow)) == 1) {
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > (alignment.Position, alignment));
}
else
break;
}
////Get next alignment
IsNextAlignment = reader.GetNextAlignment(alignment);
}
//cerr << "Done: Gather aligmenets that lie (fully or partially) within the window frame and group INDELs if there are ... " << endl;
//// Detector Corner
bool ToRealign = MeetIndelDetectorThresholdv(AlGroups);
cerr << "MeetIndelDetectorThresholdv(AlGroups).size()= " << AlGroups.size() << endl;
// **************
if (ToRealign)
{
//cerr << " ToRealign: " << refdataIter->RefName << "\t" << reflength << "\t" << winstartpos << "\t" << winendpos << "\t" << AlGroups.size() << endl;
//cerr << " minmaxRefSeqPos.at(1)= " << minmaxRefSeqPos.at(1) << " minmaxRefSeqPos.at(0)= " << minmaxRefSeqPos.at(0) << endl;
////// Perform Realign routines
int TotalAlR = 0; // Total number of alignments to be realigned
int NumAlR = 0; // Now many alignments are aligned
TotalWindowDetected++;
cerr << "#Start: Meet Threshold, Realigning ... " << endl;
if (minmaxRefSeqPos.at(1) < winendpos)
minmaxRefSeqPos.at(1) = winendpos;
if (minmaxRefSeqPos.at(0) > winstartpos)
minmaxRefSeqPos.at(0) = winstartpos;
bool IsToRealign = RealignFunction.PruningByNaiveSelectionProcedureAndConstructHaplotypes2(winstartpos, winendpos, refid, refname, minmaxRefSeqPos, reference);
if (IsToRealign == true)
{
RealignFunction.SelectHaplotypeCandidates_SmithWatermanBSv(AlGroups, minmaxRefSeqPos, SetLowComplexityRegionSWGapExt);
minmaxRefSeqPos.at(0) = -1;
minmaxRefSeqPos.at(1) = 0;
int nextwinstartpos = winendpos + 1;
int nextwinendpos = winstartpos + windowlen - 1;
if (nextwinendpos > reflength)
nextwinendpos = reflength;
//cerr << " Before Realign : " << SortRealignedAlignmentsMultimap.size() << endl;
RealignFunction.AdjustCigarsWRTChosenMultipleHaplotypesAndPrepareAlignmentsTobeWrittenOut(AlGroups, SortRealignedAlignmentsMultimap, reference, RefIDRedName, minmaxRefSeqPos, nextwinstartpos, nextwinendpos, minbaseQ, TotalAlR, NumAlR, ploidy);
IsPassDetectorNoRealignment = false; // Set flag to false to deactivate write functions
//cerr << " After Realign : " << SortRealignedAlignmentsMultimap.size() << endl;
TotalReadsAligned += NumAlR;
if (NumAlR > 0) // Realignment done
windowrealigned++;
} else
cerr << "#Done: Meet Threshold, Realigning ... " << endl;
}
if (NewReadMappedcount > 0)
TotalWindow++;
RealignFunction.Clear();
//// Move the window frame
winstartpos = winendpos + 1;
winendpos = winstartpos + windowlen - 1;
}
//// Save and Erase remaining alignments that are outside of window (Deque?)
if ((!AlGroups.empty())) {
cerr << "#Start: Write Remaining alignments and delete all alignments" << endl;
for (vector<SalRealignInfo>::iterator Iter = AlGroups.begin(); Iter != AlGroups.end(); ++Iter) {
//cerr << " Remain alignment start: " << (*Iter).al.Name << " " << Iter->al.Position << " < " << winstartpos << " " << winendpos << endl;
SortRealignedAlignmentsMultimap.insert(pair<int, BamAlignment > ((*Iter).al.Position, (*Iter).al));
}
cerr << "#Done: Write Remaining alignments and delete all alignments" << endl;
}
AlGroups.clear();
// Write Sorted remaining Alignments that are outside of window
if (!SortRealignedAlignmentsMultimap.empty())
{
for (multimap<int, BamAlignment>::iterator sraIter = SortRealignedAlignmentsMultimap.begin(); sraIter != SortRealignedAlignmentsMultimap.end(); ++sraIter)
{
//writer.SaveAlignment((*sraIter).second);
if (!writer.SaveAlignment((*sraIter).second))
cerr << writer.GetErrorString() << endl;
}
SortRealignedAlignmentsMultimap.clear();
}
}
++regionListIter;
if ((*regionListIter).LeftRefID > refid)
nextChrName = true;
}
//// If End of the chromosome position
//// increament iterator
++refdataIter;
++refid;
}
reader.Close();
writer.Close();
cerr << "##-Completed- " << endl;
cerr << " Total Reads processed = " << TotalReads << endl;
cerr << " Total Reads Aligned = " << TotalReadsAligned << endl;
cerr << " Total Window processed = " << TotalWindow << endl;
cerr << " Total Window Detected = " << TotalWindowDetected << endl;
cerr << " Total Windows Aligned = " << windowrealigned << endl;
// Restore cerr's stream buffer before terminating
if (cerrlog.is_open())
cerr.rdbuf(cerrsave);
commandline.clear();
return 0;
}
//{{{bool sort_inter_chrom_bam(string in_file_name,
bool sort_inter_chrom_bam(string in_file_name,
string out_file_name)
{
// open input BAM file
BamReader reader;
if ( !reader.Open(in_file_name) ) {
cerr << "sort ERROR: could not open " <<
in_file_name << " for reading... Aborting." << endl;
return false;
}
SamHeader header = reader.GetHeader();
if ( !header.HasVersion() )
header.Version = Constants::SAM_CURRENT_VERSION;
string header_text = header.ToString();
RefVector ref = reader.GetReferenceData();
// set up alignments buffer
BamAlignment al;
vector<BamAlignment> buffer;
buffer.reserve( (size_t)(SORT_DEFAULT_MAX_BUFFER_COUNT*1.1) );
bool bufferFull = false;
int buff_count = 0;
// iterate through file
while ( reader.GetNextAlignment(al)) {
// check buffer's usage
bufferFull = ( buffer.size() >= SORT_DEFAULT_MAX_BUFFER_COUNT );
// store alignments until buffer is "full"
if ( !bufferFull )
buffer.push_back(al);
// if buffer is "full"
else {
// so create a sorted temp file with current buffer contents
// then push "al" into fresh buffer
create_sorted_temp_file(buffer,
out_file_name,
buff_count,
header_text,
ref);
++buff_count;
buffer.push_back(al);
}
}
// handle any leftover buffer contents
if ( !buffer.empty() ) {
create_sorted_temp_file(buffer,
out_file_name,
buff_count,
header_text,
ref);
++buff_count;
}
reader.Close();
return merge_sorted_files(out_file_name, buff_count, header_text, ref);
/*
for (int i = 0; i < buff_count; ++i) {
stringstream temp_name;
temp_name << out_file_name << i;
}
*/
}
void BedIntersectPE::IntersectBamPE(string bamFile) {
// load the "B" bed file into a map so
// that we can easily compare "A" to it for overlaps
_bedB->loadBedFileIntoMap();
// open the BAM file
BamReader reader;
BamWriter writer;
reader.Open(bamFile);
// get header & reference information
string bamHeader = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// open a BAM output to stdout if we are writing BAM
if (_bamOutput == true) {
// set compression mode
BamWriter::CompressionMode compressionMode = BamWriter::Compressed;
if ( _isUncompressedBam ) compressionMode = BamWriter::Uncompressed;
writer.SetCompressionMode(compressionMode);
// open our BAM writer
writer.Open("stdout", bamHeader, refs);
}
// track the previous and current sequence
// names so that we can identify blocks of
// alignments for a given read ID.
string prevName, currName;
prevName = currName = "";
vector<BamAlignment> alignments; // vector of BAM alignments for a given ID in a BAM file.
alignments.reserve(100);
_bedA->bedType = 10; // it's a full BEDPE given it's BAM
// rip through the BAM file and convert each mapped entry to BEDPE
BamAlignment bam1, bam2;
while (reader.GetNextAlignment(bam1)) {
reader.GetNextAlignment(bam2);
if (bam1.Name != bam2.Name) {
while (bam1.Name != bam2.Name)
{
if (bam1.IsPaired())
{
cerr << "*****WARNING: Query " << bam1.Name
<< " is marked as paired, but it's mate does not occur"
<< " next to it in your BAM file. Skipping. " << endl;
}
bam1 = bam2;
reader.GetNextAlignment(bam2);
}
}
else if (bam1.IsPaired() && bam1.IsPaired()) {
ProcessBamBlock(bam1, bam2, refs, writer);
}
}
// close up
reader.Close();
if (_bamOutput == true) {
writer.Close();
}
}
int main (int argc, char *argv[]) {
bool produceUnCompressedBAM=false;
bool verbose=false;
bool ancientDNA=false;
bool keepOrig=false;
string adapter_F=options_adapter_F_BAM;
string adapter_S=options_adapter_S_BAM;
string adapter_chimera=options_adapter_chimera_BAM;
string key="";
bool allowMissing=false;
int trimCutoff=1;
bool allowAligned=false;
bool printLog=false;
string logFileName;
BamReader reader;
BamWriter writer;
string bamFile;
string bamFileOUT="";
string key1;
string key2;
bool useDist=false;
double location=-1.0;
double scale =-1.0;
bool fastqFormat=false;
string fastqfile1 = "";
string fastqfile2 = "";
string fastqoutfile = "";
bool singleEndModeFQ=true;
const string usage=string(string(argv[0])+
" [options] BAMfile"+"\n"+
"\nThis program takes an unaligned BAM where mates are consecutive\nor fastq files and trims and merges reads\n"+
"\n\tYou can specify a unaligned bam file or one or two fastq :\n"+
"\t\t"+"-fq1" +"\t\t"+"First fastq"+"\n"+
"\t\t"+"-fq2" +"\t\t"+"Second fastq file (for paired-end)"+"\n"+
"\t\t"+"-fqo" +"\t\t"+"Output fastq prefix"+"\n\n"+
//"\t"+"-p , --PIPE"+"\n\t\t"+"Read BAM from and write it to PIPE"+"\n"+
"\t"+"-o , --outfile" +"\t\t"+"Output (BAM format)."+"\n"+
"\t"+"-u " +"\t\t"+"Produce uncompressed bam (good for pipe)"+"\n"+
// "\t"+" , --outprefix" +"\n\t\t"+"Prefix for output files (default '"+outprefix+"')."+"\n"+
//"\t"+" , --SAM" +"\n\t\t"+"Output SAM not BAM."+"\n"+
"\t"+"--aligned" +"\t\t"+"Allow reads to be aligned (default "+boolStringify(allowAligned)+")"+"\n"+
"\t"+"-v , --verbose" +"\t\t"+"Turn all messages on (default "+boolStringify(verbose)+")"+"\n"+
"\t"+"--log [log file]" +"\t"+"Print a tally of merged reads to this log file (default only to stderr)"+"\n"+
"\n\t"+"Paired End merging/Single Read trimming options"+"\n"+
"\t\t"+"You can specify either:"+"\n"+
"\t\t\t"+"--ancientdna"+"\t\t\t"+"ancient DNA (default "+boolStringify(ancientDNA)+")"+"\n"+
"\t\t"+" "+"\t\t\t\t"+"this allows for partial overlap"+"\n"+
"\n\t\t"+"or if you know your size length distribution:"+"\n"+
"\t\t\t"+"--loc"+"\t\t\t\t"+"Location for lognormal dist. (default none)"+"\n"+
"\t\t\t"+"--scale"+"\t\t\t\t"+"Scale for lognormal dist. (default none)"+"\n"+
// "\t\t\t\t\t\t\tGood for merging ancient DNA reads into a single sequence\n\n"
"\n\t\t"+"--keepOrig"+"\t\t\t\t"+"Write original reads if they are trimmed or merged (default "+boolStringify(keepOrig)+")"+"\n"+
"\t\t\t\t\t\t\tSuch reads will be marked as PCR duplicates\n\n"
"\t\t"+"-f , --adapterFirstRead" +"\t\t\t"+"Adapter that is observed after the forward read (def. Multiplex: "+options_adapter_F_BAM.substr(0,30)+")"+"\n"+
"\t\t"+"-s , --adapterSecondRead" +"\t\t"+"Adapter that is observed after the reverse read (def. Multiplex: "+options_adapter_S_BAM.substr(0,30)+")"+"\n"+
"\t\t"+"-c , --FirstReadChimeraFilter" +"\t\t"+"If the forward read looks like this sequence, the cluster is filtered out.\n\t\t\t\t\t\t\tProvide several sequences separated by comma (def. Multiplex: "+options_adapter_chimera_BAM.substr(0,30)+")"+"\n"+
"\t\t"+"-k , --key"+"\t\t\t\t"+"Key sequence with which each sequence starts. Comma separate for forward and reverse reads. (default '"+key+"')"+"\n"+
"\t\t"+"-i , --allowMissing"+"\t\t\t"+"Allow one base in one key to be missing or wrong. (default "+boolStringify(allowMissing)+")"+"\n"+
"\t\t"+"-t , --trimCutoff"+"\t\t\t"+"Lowest number of adapter bases to be observed for single Read trimming (default "+stringify(trimCutoff)+")");
if( (argc== 1) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cout<<"Usage:"<<endl;
cout<<""<<endl;
cout<<usage<<endl;
return 1;
}
for(int i=1;i<(argc-1);i++){ //all but the last arg
if(strcmp(argv[i],"-fq1") == 0 ){
fastqfile1=string(argv[i+1]);
fastqFormat=true;
i++;
continue;
}
if(strcmp(argv[i],"-fq2") == 0 ){
fastqfile2=string(argv[i+1]);
fastqFormat=true;
singleEndModeFQ=false;
i++;
continue;
}
if(strcmp(argv[i],"-fqo") == 0 ){
fastqoutfile=string(argv[i+1]);
fastqFormat=true;
i++;
continue;
}
if(strcmp(argv[i],"--log") == 0 ){
logFileName =string(argv[i+1]);
printLog=true;
i++;
continue;
}
if(strcmp(argv[i],"-p") == 0 || strcmp(argv[i],"--PIPE") == 0 ){
cerr<<"This version no longer works with pipe, exiting"<<endl;
return 1;
}
if(strcmp(argv[i],"-u") == 0 ){
produceUnCompressedBAM=true;
continue;
}
if(strcmp(argv[i],"--aligned") == 0 ){
allowAligned=true;
continue;
}
if(strcmp(argv[i],"-o") == 0 || strcmp(argv[i],"--outfile") == 0 ){
bamFileOUT =string(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-v") == 0 || strcmp(argv[i],"--verbose") == 0 ){
verbose=true;
continue;
}
if(strcmp(argv[i],"--ancientdna") == 0 ){
ancientDNA=true;
continue;
}
if(strcmp(argv[i],"--keepOrig") == 0 ){
keepOrig=true;
continue;
}
if(strcmp(argv[i],"--loc") == 0 ){
location =destringify<double>(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"--scale") == 0 ){
scale =destringify<double>(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-f") == 0 || strcmp(argv[i],"--adapterFirstRead") == 0 ){
adapter_F =string(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-s") == 0 || strcmp(argv[i],"--adapterSecondRead") == 0 ){
adapter_S =string(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-c") == 0 || strcmp(argv[i],"--FirstReadChimeraFilter") == 0 ){
adapter_chimera =string(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-k") == 0 || strcmp(argv[i],"--keys") == 0 ){
key =string(argv[i+1]);
i++;
continue;
}
if(strcmp(argv[i],"-i") == 0 || strcmp(argv[i],"--allowMissing") == 0 ){
allowMissing=true;
continue;
}
if(strcmp(argv[i],"-t") == 0 || strcmp(argv[i],"--trimCutoff") == 0 ){
trimCutoff=atoi(argv[i+1]);
i++;
continue;
}
cerr<<"Unknown option "<<argv[i] <<" exiting"<<endl;
return 1;
}
bamFile=argv[argc-1];
if( (location != -1.0 && scale == -1.0) ||
(location == -1.0 && scale != -1.0) ){
cerr<<"Cannot specify --location without specifying --scale"<<endl;
return 1;
}
if( (location != -1.0 && scale != -1.0) ){
useDist=true;
if(ancientDNA){
cerr<<"Cannot specify --location/--scale and --ancientDNA"<<endl;
return 1;
}
}
MergeTrimReads mtr (adapter_F,adapter_S,adapter_chimera,
key1,key2,
trimCutoff,allowMissing,ancientDNA,location,scale,useDist);
fqwriters onereadgroup;
if(fastqFormat){
if( bamFileOUT != "" || produceUnCompressedBAM || allowAligned){
cerr<<"ERROR : Cannot specify options like -o, -u or --allowAligned for fastq"<<endl;
return 1;
}
if(fastqfile1 == ""){
cerr<<"ERROR : Must specify as least the first file for fastq"<<endl;
return 1;
}
FastQParser * fqp1;
FastQParser * fqp2;
if(singleEndModeFQ){
fqp1 = new FastQParser (fastqfile1);
string outdirs = fastqoutfile+".fq.gz";
string outdirsf = fastqoutfile+".fail.fq.gz";
onereadgroup.single.open(outdirs.c_str(), ios::out);
onereadgroup.singlef.open(outdirsf.c_str(), ios::out);
if(!onereadgroup.single.good()){ cerr<<"Cannot write to file "<<outdirs<<endl; return 1; }
if(!onereadgroup.singlef.good()){ cerr<<"Cannot write to file "<<outdirsf<<endl; return 1; }
}else{
fqp1 = new FastQParser (fastqfile1);
fqp2 = new FastQParser (fastqfile2);
string outdirs = fastqoutfile+".fq.gz";
string outdir1 = fastqoutfile+"_r1.fq.gz";
string outdir2 = fastqoutfile+"_r2.fq.gz";
string outdirsf = fastqoutfile+".fail.fq.gz";
string outdir1f = fastqoutfile+"_r1.fail.fq.gz";
string outdir2f = fastqoutfile+"_r2.fail.fq.gz";
onereadgroup.single.open(outdirs.c_str(), ios::out);
onereadgroup.pairr1.open(outdir1.c_str(), ios::out);
onereadgroup.pairr2.open(outdir2.c_str(), ios::out);
onereadgroup.singlef.open(outdirsf.c_str(), ios::out);
onereadgroup.pairr1f.open(outdir1f.c_str(), ios::out);
onereadgroup.pairr2f.open(outdir2f.c_str(), ios::out);
if(!onereadgroup.single.good()){ cerr<<"Cannot write to file "<<outdirs<<endl; return 1; }
if(!onereadgroup.pairr1.good()){ cerr<<"Cannot write to file "<<outdir1<<endl; return 1; }
if(!onereadgroup.pairr2.good()){ cerr<<"Cannot write to file "<<outdir2<<endl; return 1; }
if(!onereadgroup.singlef.good()){ cerr<<"Cannot write to file "<<outdirsf<<endl; return 1; }
if(!onereadgroup.pairr1f.good()){ cerr<<"Cannot write to file "<<outdir1f<<endl; return 1; }
if(!onereadgroup.pairr2f.good()){ cerr<<"Cannot write to file "<<outdir2f<<endl; return 1; }
}
unsigned int totalSeqs=0;
while(fqp1->hasData()){
FastQObj * fo1=fqp1->getData();
vector<string> def1=allTokens( *(fo1->getID()), ' ' );
string def1s=def1[0];
FastQObj * fo2;
string def2s;
string ext2s;
if(!singleEndModeFQ){
if(!fqp2->hasData()){
cerr << "ERROR: Discrepency between fastq files at record " << *(fo1->getID()) <<endl;
return 1;
}
fo2=fqp2->getData();
vector<string> def2=allTokens( *(fo2->getID()), ' ' );
def2s=def2[0];
if(strEndsWith(def1s,"/1")){
def1s=def1s.substr(0,def1s.size()-2);
}
if(strEndsWith(def2s,"/2")){
def2s=def2s.substr(0,def2s.size()-2);
}
if(strBeginsWith(def1s,"@")){
def1s=def1s.substr(1,def1s.size()-1);
}
if(strBeginsWith(def2s,"@")){
def2s=def2s.substr(1,def2s.size()-1);
}
if(def1s != def2s){
cerr << "ERROR: Discrepency between fastq files, different names " << *(fo1->getID()) <<" and "<< *(fo2->getID()) <<endl;
return 1;
}
merged result= mtr.process_PE(*(fo1->getSeq()),*(fo1->getQual()),
*(fo2->getSeq()),*(fo2->getQual()));
mtr.incrementCountall();
if(result.code != ' '){ //keys or chimeras
if(result.code == 'K'){
mtr.incrementCountfkey();
}else{
if(result.code == 'D'){
mtr.incrementCountchimera();
}else{
cerr << "leehom: Wrong return code =\""<<result.code<<"\""<<endl;
exit(1);
}
}
onereadgroup.pairr2f<<"@"<<def2s<<"/2" <<endl <<*(fo2->getSeq())<<endl<<"+"<<endl <<*(fo2->getQual())<<endl;
onereadgroup.pairr1f<<"@"<<def1s<<"/1" <<endl <<*(fo1->getSeq())<<endl<<"+"<<endl <<*(fo1->getQual())<<endl;
continue;
}else{
if(result.sequence != ""){ //new sequence
onereadgroup.single<<"@"<<def1s<<"" <<endl << result.sequence<<endl<<"+"<<endl <<result.quality<<endl;
if( result.sequence.length() > max(fo1->getSeq()->length(),fo2->getSeq()->length()) ){
mtr.incrementCountmergedoverlap();
}else{
mtr.incrementCountmerged();
}
}else{ //keep as is
mtr.incrementCountnothing();
onereadgroup.pairr2<<"@"<<def2s<<"/2" <<endl <<*(fo2->getSeq())<<endl<<"+"<<endl <<*(fo2->getQual())<<endl;
onereadgroup.pairr1<<"@"<<def1s<<"/1" <<endl <<*(fo1->getSeq())<<endl<<"+"<<endl <<*(fo1->getQual())<<endl;
}
}
}else{
merged result=mtr.process_SR(*(fo1->getSeq()),*(fo1->getQual()));
mtr.incrementCountall();
if(result.code != ' '){ //either chimera or missing key
if(result.code == 'K'){
mtr.incrementCountfkey();
}else{
if(result.code == 'D'){
mtr.incrementCountchimera();
}else{
cerr << "leehom: Wrong return code =\""<<result.code<<"\""<<endl;
exit(1);
}
}
onereadgroup.singlef<<""<<*(fo1->getID())<<"" <<endl << *(fo1->getSeq())<<endl<<"+"<<endl <<*(fo1->getQual())<<endl;
continue;
}
if(result.sequence != ""){ //new sequence
mtr.incrementCounttrimmed();
onereadgroup.single<<""<<*(fo1->getID())<<"" <<endl << result.sequence<<endl<<"+"<<endl <<result.quality<<endl;
}else{
mtr.incrementCountnothing();
onereadgroup.single<<""<<*(fo1->getID())<<"" <<endl << *(fo1->getSeq())<<endl<<"+"<<endl <<*(fo1->getQual())<<endl;
}
}
totalSeqs++;
}
delete fqp1;
if(!singleEndModeFQ){
delete fqp2;
}
if(singleEndModeFQ){
onereadgroup.single.close();
onereadgroup.singlef.close();
}else{
onereadgroup.single.close();
onereadgroup.pairr1.close();
onereadgroup.pairr2.close();
onereadgroup.singlef.close();
onereadgroup.pairr1f.close();
onereadgroup.pairr2f.close();
}
//fastq
}else{
//else BAM
// initMerge();
// set_adapter_sequences(adapter_F,
// adapter_S,
// adapter_chimera);
// set_options(trimCutoff,allowMissing,mergeoverlap);
if(key != ""){
size_t found=key.find(",");
if (found == string::npos){ //single end reads
key1=key;
key2="";
} else{ //paired-end
key1=key.substr(0,found);
key2=key.substr(found+1,key.length()-found+1);
}
}
if( bamFileOUT == "" ){
cerr<<"The output must be a be specified, exiting"<<endl;
return 1;
}
if ( !reader.Open(bamFile) ) {
cerr << "Could not open input BAM file "<<bamFile << endl;
return 1;
}
SamHeader header = reader.GetHeader();
string pID = "mergeTrimReadsBAM";
string pName = "mergeTrimReadsBAM";
string pCommandLine = "";
for(int i=0;i<(argc);i++){
pCommandLine += (string(argv[i])+" ");
}
putProgramInHeader(&header,pID,pName,pCommandLine,returnGitHubVersion(string(argv[0]),".."));
const RefVector references = reader.GetReferenceData();
//we will not call bgzip with full compression, good for piping into another program to
//lessen the load on the CPU
if(produceUnCompressedBAM)
writer.SetCompressionMode(BamWriter::Uncompressed);
if ( !writer.Open(bamFileOUT,header,references) ) {
cerr << "Could not open output BAM file "<<bamFileOUT << endl;
return 1;
}
SamHeader sh=reader.GetHeader();
//Up to the user to be sure that a sequence is followed by his mate
// if(!sh.HasSortOrder() ||
// sh.SortOrder != "queryname"){
// cerr << "Bamfile must be sorted by queryname" << endl;
// return 1;
// }
BamAlignment al;
BamAlignment al2;
bool al2Null=true;
while ( reader.GetNextAlignment(al) ) {
if(al.IsMapped() || al.HasTag("NM") || al.HasTag("MD") ){
if(!allowAligned){
cerr << "Reads should not be aligned" << endl;
return 1;
}else{
//should we remove tags ?
}
}
if(al.IsPaired() &&
al2Null ){
al2=al;
al2Null=false;
continue;
}else{
if(al.IsPaired() &&
!al2Null){
bool result = mtr.processPair(al,al2);
if( result ){//was merged
BamAlignment orig;
BamAlignment orig2;
if(keepOrig){
orig2 = al2;
orig = al;
}
writer.SaveAlignment(al);
if(keepOrig){
orig.SetIsDuplicate(true);
orig2.SetIsDuplicate(true);
writer.SaveAlignment(orig2);
writer.SaveAlignment(orig);
}
//the second record is empty
}else{
//keep the sequences as pairs
writer.SaveAlignment(al2);
writer.SaveAlignment(al);
}
//
// SINGLE END
//
}else{
BamAlignment orig;
if(keepOrig){
orig =al;
}
mtr.processSingle(al);
if(keepOrig){
//write duplicate
if(orig.QueryBases.length() != al.QueryBases.length()){
orig.SetIsDuplicate(true);
writer.SaveAlignment(orig);
}
}
writer.SaveAlignment(al);
} //end single end
al2Null=true;
}//second pair
} //while al
reader.Close();
writer.Close();
} //else BAM
cerr <<mtr.reportSingleLine()<<endl;
if(printLog){
ofstream fileLog;
fileLog.open(logFileName.c_str());
if (fileLog.is_open()){
fileLog <<mtr.reportMultipleLines() <<endl;
}else{
cerr << "Unable to print to file "<<logFileName<<endl;
}
fileLog.close();
}
return 0;
}
int main(const int argc, char* const argv[]) {
int c, min_mapQ=0, seed=chrono::system_clock::now().time_since_epoch().count();
unsigned int flag_on=0, flag_off=0;
string fn_tgt, fn_in, fn_out="", out_format="b";
while ((c = getopt(argc, argv, "SbBcCt:h1Ho:q:f:F:ul:r:?T:R:L:s:@:m:x:U:")) >= 0) {
switch (c) {
case 's': seed = atoi(optarg); break;
case 'm': break;
case 'c': break;
case 'S': break;
case 'b': break;
case 'C': break;
case 'h': break;
case 'H': break;
case 'o': fn_out = optarg; break;
case 'U': break;
case 'f': flag_on |= strtol(optarg, 0, 0); break;
case 'F': flag_off |= strtol(optarg, 0, 0); break;
case 'q': min_mapQ = atoi(optarg); break;
case 'u': out_format = "u"; break;
case '1': break;
case 'l': break;
case 'r': break;
case 't': fn_tgt = optarg; break;
case 'R': break;
case '?': return usage();
case 'T': break;
case 'B': break;
case '@': break;
case 'x': break;
default: return usage();
}
}
if (fn_tgt.compare("") == 0) return usage();
if (argc == optind) return usage();
fn_in = argv[optind];
BamReader reader;
if (!reader.Open(fn_in)) {
cerr << "ERROR: cannot open [" << fn_in << "] for reading\n";
return 1;
}
if (!reader.LocateIndex()) {
cerr << "ERROR: cannot find BAM index for [" << fn_in << "]\n";
return 1;
}
const SamHeader header = reader.GetHeader();
if (header.SortOrder.compare("coordinate") != 0) {
cerr << "ERROR: [" << fn_in << "] not sorted by coordinate\n";
return 1;
}
const RefVector refseq = reader.GetReferenceData();
vector<BamRegion> regions;
vector<unsigned int> src_depths, tgt_depths;
if (read_region_depth(fn_tgt.c_str(), reader, regions, src_depths, tgt_depths) != 0) return 1;
BamWriter writer;
if (!writer.Open(fn_out, header, refseq)) {
cerr << "ERROR: cannot open [" << fn_out << "] for writing\n";
return 1;
}
BamAlignment aln;
vector<BamAlignment> reads;
vector<string> paired, unpaired;
unordered_map<int, int> kept;
unordered_map<string, unsigned int> seen, sampled;
unordered_map<string, vector<int> > pool;
for (size_t i=0; i<regions.size(); ++i) {
reads.clear();
paired.clear();
unpaired.clear();
kept.clear();
pool.clear();
char region_string[256];
sprintf(region_string, "%s:%d-%d", refseq[regions[i].LeftRefID].RefName.c_str(), regions[i].LeftPosition, regions[i].RightPosition);
if (!reader.SetRegion(regions[i])) {
cerr << "WARNING: failed to locate [" << region_string << "]\n";
//cerr << "WARNING: failed to locate [" << refseq[regions[i].LeftRefID].RefName << ':' << regions[i].LeftPosition << '-' << regions[i].RightPosition << "]\n";
continue;
}
while (reader.GetNextAlignment(aln)) {
if ((aln.AlignmentFlag & flag_on) == flag_on && !(aln.AlignmentFlag & flag_off) && aln.MapQuality >= min_mapQ)
reads.push_back(aln);
}
if (reads.size() == 0) continue;
unsigned int depth = 0;
for (size_t k=0; k<reads.size(); ++k) {
aln = reads[k];
string rn = aln.Name;
if (seen.find(rn) != seen.end()) { // if seen in previous regions
if (sampled.find(rn) != sampled.end()) { // if self or mate sampled before, sample it
if (sampled[rn] != aln.AlignmentFlag) kept[k] = 1; // if mate sampled before, keep it
depth += get_overlap(aln, regions[i]);
}
if (seen[rn] != aln.AlignmentFlag) seen[rn] = aln.AlignmentFlag;
}
else { // if not seen in previous regions
pool[rn].push_back(k);
}
if (depth > tgt_depths[i]) break;
}
if (depth < tgt_depths[i]) {
for (auto it=pool.begin(); it!=pool.end(); ++it) {
if (it->second.size()>1)
paired.push_back(it->first);
else
unpaired.push_back(it->first);
}
shuffle(paired.begin(), paired.end(), default_random_engine(seed));
shuffle(unpaired.begin(), unpaired.end(), default_random_engine(seed));
int n1=paired.size(), n2=unpaired.size(), k1, k2, k3;
while (depth < tgt_depths[i] && n1+n2 > 0) {
if (n1>0) {
k1 = pool[paired[--n1]][0];
k2 = pool[paired[n1]][1];
depth += get_overlap(reads[k1], regions[i]);
depth += get_overlap(reads[k2], regions[i]);
kept[k1] = 1; kept[k2] = 1;
continue;
}
if (n2>0) {
k3 = pool[unpaired[--n2]][0];
depth += get_overlap(reads[k3], regions[i]);
kept[k3] = 1;
continue;
}
}
}
for (auto it=pool.begin(); it!=pool.end(); ++it) {
string rn = it->first;
seen[rn] = reads[pool[rn].back()].AlignmentFlag;
}
for (auto it=kept.begin(); it!=kept.end(); ++it) {
int k = it->first;
string rn = reads[k].Name;
sampled[rn] = reads[k].AlignmentFlag;
writer.SaveAlignment(reads[k]);
}
cerr << "INFO: target=[" << tgt_depths[i] << "], actual=[" << depth << "], N(reads)=[" << reads.size() << "], N(kept)=[" << kept.size() << "] at [" << region_string << "]\n";
}
reader.Close();
return 0;
}
// generates mutiple sorted temp BAM files from single unsorted BAM file
bool SortTool::SortToolPrivate::GenerateSortedRuns(void) {
// open input BAM file
BamReader reader;
if ( !reader.Open(m_settings->InputBamFilename) ) {
cerr << "bamtools sort ERROR: could not open " << m_settings->InputBamFilename
<< " for reading... Aborting." << endl;
return false;
}
// get basic data that will be shared by all temp/output files
SamHeader header = reader.GetHeader();
header.SortOrder = ( m_settings->IsSortingByName
? Constants::SAM_HD_SORTORDER_QUERYNAME
: Constants::SAM_HD_SORTORDER_COORDINATE );
m_headerText = header.ToString();
m_references = reader.GetReferenceData();
// set up alignments buffer
BamAlignment al;
vector<BamAlignment> buffer;
buffer.reserve( (size_t)(m_settings->MaxBufferCount*1.1) );
bool bufferFull = false;
// if sorting by name, we need to generate full char data
// so can't use GetNextAlignmentCore()
if ( m_settings->IsSortingByName ) {
// iterate through file
while ( reader.GetNextAlignment(al)) {
// check buffer's usage
bufferFull = ( buffer.size() >= m_settings->MaxBufferCount );
// store alignments until buffer is "full"
if ( !bufferFull )
buffer.push_back(al);
// if buffer is "full"
else {
// push any unmapped reads into buffer,
// don't want to split these into a separate temp file
if ( !al.IsMapped() )
buffer.push_back(al);
// "al" is mapped, so create a sorted temp file with current buffer contents
// then push "al" into fresh buffer
else {
CreateSortedTempFile(buffer);
buffer.push_back(al);
}
}
}
}
// sorting by position, can take advantage of GNACore() speedup
else {
// iterate through file
while ( reader.GetNextAlignmentCore(al) ) {
// check buffer's usage
bufferFull = ( buffer.size() >= m_settings->MaxBufferCount );
// store alignments until buffer is "full"
if ( !bufferFull )
buffer.push_back(al);
// if buffer is "full"
else {
// push any unmapped reads into buffer,
// don't want to split these into a separate temp file
if ( !al.IsMapped() )
buffer.push_back(al);
// "al" is mapped, so create a sorted temp file with current buffer contents
// then push "al" into fresh buffer
else {
CreateSortedTempFile(buffer);
buffer.push_back(al);
}
}
}
}
// handle any leftover buffer contents
if ( !buffer.empty() )
CreateSortedTempFile(buffer);
// close reader & return success
reader.Close();
return true;
}
void TagBam::Tag() {
// open the annotations files for processing;
OpenAnnoFiles();
// open the BAM file
BamReader reader;
BamWriter writer;
if (!reader.Open(_bamFile)) {
cerr << "Failed to open BAM file " << _bamFile << endl;
exit(1);
}
// get header & reference information
string bamHeader = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// set compression mode
BamWriter::CompressionMode compressionMode = BamWriter::Compressed;
// if ( _isUncompressedBam ) compressionMode = BamWriter::Uncompressed;
writer.SetCompressionMode(compressionMode);
// open our BAM writer
writer.Open("stdout", bamHeader, refs);
// rip through the BAM file and test for overlaps with each annotation file.
BamAlignment al;
vector<BED> hits;
while (reader.GetNextAlignment(al)) {
if (al.IsMapped() == true) {
BED a;
a.chrom = refs.at(al.RefID).RefName;
a.start = al.Position;
a.end = al.GetEndPosition(false, false);
a.strand = "+";
if (al.IsReverseStrand()) a.strand = "-";
ostringstream annotations;
// annotate the BAM file based on overlaps with the annotation files.
for (size_t i = 0; i < _annoFiles.size(); ++i)
{
// grab the current annotation file.
BedFile *anno = _annoFiles[i];
if (!_useNames && !_useScores && !_useIntervals) {
// add the label for this annotation file to tag if there is overlap
if (anno->anyHits(a.chrom, a.start, a.end, a.strand,
_sameStrand, _diffStrand, _overlapFraction, false))
{
annotations << _annoLabels[i] << ";";
}
}
// use the score field
else if (!_useNames && _useScores && !_useIntervals) {
anno->allHits(a.chrom, a.start, a.end, a.strand,
hits, _sameStrand, _diffStrand, 0.0, false);
for (size_t i = 0; i < hits.size(); ++i) {
annotations << hits[i].score;
if (i < hits.size() - 1) annotations << ",";
}
if (hits.size() > 0) annotations << ";";
hits.clear();
}
// use the name field from the annotation files to populate tag
else if (_useNames && !_useScores && !_useIntervals) {
anno->allHits(a.chrom, a.start, a.end, a.strand,
hits, _sameStrand, _diffStrand, 0.0, false);
for (size_t j = 0; j < hits.size(); ++j) {
annotations << hits[j].name;
if (j < hits.size() - 1) annotations << ",";
}
if (hits.size() > 0) annotations << ";";
hits.clear();
}
// use the full interval information annotation files to populate tag
else if (!_useNames && !_useScores && _useIntervals) {
anno->allHits(a.chrom, a.start, a.end, a.strand,
hits, _sameStrand, _diffStrand, 0.0, false);
for (size_t j = 0; j < hits.size(); ++j) {
annotations << _annoLabels[i] << ":" <<
hits[j].chrom << ":" <<
hits[j].start << "-" <<
hits[j].end << "," <<
hits[j].name << "," <<
hits[j].score << "," <<
hits[j].strand;
if (j < hits.size() - 1) annotations << ",";
}
if (hits.size() > 0) annotations << ";";
hits.clear();
}
}
// were there any overlaps with which to make a tag?
if (annotations.str().size() > 0) {
al.AddTag(_tag, "Z", annotations.str().substr(0, annotations.str().size() - 1)); // get rid of the last ";"
}
}
writer.SaveAlignment(al);
}
reader.Close();
writer.Close();
// close the annotations files;
CloseAnnoFiles();
}
int
main(int argc, char* argv[]) {
// validate argument count
if( argc != 2 ) {
cerr << "USAGE: " << argv[0] << " <input BAM file> " << endl;
return EXIT_FAILURE;
}
string filename = argv[1];
//cerr << "Printing alignments from file: " << filename << endl;
BamReader reader;
if (!reader.Open(filename)) {
cerr << "could not open filename " << filename << endl;
return EXIT_FAILURE;
}
cerr << filename << ": Done opening" << endl;
// Header can't be used to accurately determine sort order because samtools never
// changes it; instead, check after loading each read as is done with "samtools index"
// We don't need to load an index (right?)
// if (!reader.LocateIndex()) {
// const string index_filename = filename + ".bai";
// if (!reader.OpenIndex(index_filename)) {
// cerr << "could not open index" << endl;
// }
// }
const SamHeader header = reader.GetHeader();
cerr << filename << ": Done getting header" << endl;
const RefVector refs = reader.GetReferenceData();
cerr << filename << ": Done getting reference data" << endl;
BamWriter writer;
if (! output_bam_filename.empty()) {
if (! writer.Open(output_bam_filename, header, refs)) {
cerr << "Could not open BAM output file " << output_bam_filename << endl;
return EXIT_FAILURE;
}
cerr << filename << ": Done opening BAM output file " << output_bam_filename << endl;
}
alignmentMap read1Map; // a single map, for all reads awaiting their mate
typedef map<string,int32_t> stringMap;
typedef stringMap::iterator stringMapI;
stringMap ref_mates;
// alignmentMap read1Map, read2Map;
BamAlignment full_al;
int32_t count = 0;
uint32_t max_reads_in_map = 0;
int32_t n_reads_skipped_unmapped = 0;
int32_t n_reads_skipped_mate_unmapped = 0;
int32_t n_reads_skipped_wont_see_mate = 0;
int32_t n_reads_skipped_mate_tail_est = 0;
int32_t n_reads_skipped_ref_mate = 0;
int32_t n_reads = 0;
int32_t n_singleton_reads = 0;
int32_t last_RefID = -1;
int32_t last_Position = -1;
cerr << filename << ": Looking for up to " << pairs_to_process << " link pairs,"
<< " total tail = " << link_pair_total_tail
<< " critical tail = " << link_pair_crit_tail
<< ", must be on diff chromosome = " << link_pair_diff_chrom << endl;
while (reader.GetNextAlignment(full_al)
&& (! pairs_to_process || count < pairs_to_process)) {
BamAlignment al = full_al;
//printAlignmentInfo(al, refs);
//++count;
++n_reads;
if (last_RefID < 0) last_RefID = al.RefID;
if (last_Position < 0) last_Position = al.Position;
if (al.RefID > last_RefID) {
// We've moved to the next reference sequence
// Clean up reads with mates expected here that haven't been seen
if (debug_ref_mate) {
cerr << "MISSED " << ref_mates.size() << " ref_mates on this reference "
<< last_RefID << " " << refs[last_RefID].RefName << endl;
}
for (stringMapI rmI = ref_mates.begin(); rmI != ref_mates.end(); ++rmI) {
++n_reads_skipped_ref_mate;
read1Map.erase(read1Map.find(rmI->first));
ref_mates.erase(ref_mates.find(rmI->first));
}
last_RefID = al.RefID;
last_Position = al.Position;
} else if (! isCoordinateSorted(al.RefID, al.Position, last_RefID, last_Position)) {
cerr << filename << " is not sorted, " << al.Name << " out of position" << endl;
return EXIT_FAILURE;
}
if (! al.IsMapped()) { ++n_reads_skipped_unmapped; continue; }
if (! al.IsMateMapped()) { ++n_reads_skipped_mate_unmapped; continue; }
alignmentMapI mI = read1Map.find(al.Name);
if (mI == read1Map.end()) {
// the read name has not been seen before
if (al.MateRefID < al.RefID
|| (al.MateRefID == al.RefID && al.MatePosition < al.Position)) {
// we should have seen its mate earlier, so skip it
++n_reads_skipped_wont_see_mate;
continue;
}
// If the mate likely to also be a link pair candidate, add the read
int32_t mate_tail_est = readTailS(al.IsMateMapped(), al.IsMateReverseStrand(),
al.MatePosition, refs[al.MateRefID].RefLength, max_read_length);
if (mate_tail_est <= mate_tail_est_crit) {
// the mate tail estimate suggests it might be a link pair candidate
read1Map[al.Name] = al; // add the read to the map
} else {
// the mate tail estimate appears too long for the mate to be a candidate
++n_reads_skipped_mate_tail_est;
continue;
}
if (read1Map.size() > max_reads_in_map) max_reads_in_map = read1Map.size();
if (al.MateRefID == al.RefID && al.MatePosition >= al.Position) {
// the mate is expected later on this contig
ref_mates[al.Name] = al.MateRefID;
}
} else {
// get the mate's alignment, and process the pair
const BamAlignment& al_mate = mI->second;
if (processReadPair(al, al_mate, refs, link_pair_total_tail,
link_pair_crit_tail, link_pair_diff_chrom)) {
++count;
// write to the new BAM file, if the string is not empty
if (! output_bam_filename.empty()) {
writer.SaveAlignment(al_mate); // the first one seen
writer.SaveAlignment(al); // the second one seen
}
}
read1Map.erase(mI);
if (al.MateRefID == al.RefID) {
stringMapI rmI = ref_mates.find(al.Name);
if (rmI == ref_mates.end()) {
cerr << "expected a ref_mate, couldn't find its name: " << al.Name << endl;
return EXIT_FAILURE;
}
ref_mates.erase(rmI);
}
}
}
cerr << "===============================" << endl;
cerr << read1Map.size() << " alignments left in read1Map" << endl;
cerr << max_reads_in_map << " maximum number of reads in read1Map" << endl;
cerr << count << " pairs processed" << endl;
cerr << "===============================" << endl;
cerr << n_reads << " total reads" << endl;
cerr << n_singleton_reads << " singleton reads" << endl;
cerr << n_reads_skipped_unmapped << " reads skipped because unmapped" << endl;
cerr << n_reads_skipped_mate_unmapped << " reads skipped because mate unmapped" << endl;
cerr << n_reads_skipped_wont_see_mate << " reads skipped because mate won't be seen" << endl;
cerr << n_reads_skipped_mate_tail_est << " reads skipped because mate tail appears too long" << endl;
cerr << n_reads_skipped_ref_mate << " reads skipped because mate not on reference" << endl;
reader.Close();
if (! output_bam_filename.empty()) {
writer.Close();
}
return EXIT_SUCCESS;
}
void BamToFastq::PairedFastqUseTags() {
// open the 1st fastq file for writing
ofstream fq1(_fastq1.c_str(), ios::out);
if ( !fq1 ) {
cerr << "Error: The first fastq file (" << _fastq1 << ") could not be opened. Exiting!" << endl;
exit (1);
}
// open the 2nd fastq file for writing
ofstream fq2(_fastq2.c_str(), ios::out);
if ( !fq2 ) {
cerr << "Error: The second fastq file (" << _fastq2 << ") could not be opened. Exiting!" << endl;
exit (1);
}
// open the BAM file
BamReader reader;
reader.Open(_bamFile);
// rip through the BAM file and convert each mapped entry to BEDPE
BamAlignment bam1, bam2;
while (reader.GetNextAlignment(bam1)) {
reader.GetNextAlignment(bam2);
if (bam1.Name != bam2.Name) {
while (bam1.Name != bam2.Name)
{
if (bam1.IsPaired())
{
cerr << "*****WARNING: Query " << bam1.Name
<< " is marked as paired, but it's mate does not occur"
<< " next to it in your BAM file. Skipping. " << endl;
}
bam1 = bam2;
reader.GetNextAlignment(bam2);
}
}
else if (bam1.IsPaired() && bam2.IsPaired()) {
// assume the R2 and Q2 tags are on the + strand.
string mateSequence, mateQualities;
bam1.GetTag("R2", mateSequence);
bam1.GetTag("Q2", mateQualities);
string seq1 = bam1.QueryBases;
string qual1 = bam1.Qualities;
if (bam1.IsReverseStrand() == true) {
reverseComplement(seq1);
reverseSequence(qual1);
}
// since the info for both ends are contained in each BAM record,
// we only need to process one of the two records (bam1) in order
// to produce FASTQ entries for both ends.
// NOTE: Assumes that R2 and Q2 have already been rev
// and revcomped if necessary
if (bam1.IsFirstMate() == true) {
// end1
fq1 << "@" << bam1.Name << "/1" << endl;
fq1 << seq1 << endl;
fq1 << "+" << endl;
fq1 << qual1 << endl;
// end2
fq2 << "@" << bam1.Name << "/2" <<endl;
fq2 << mateSequence << endl;
fq2 << "+" << endl;
fq2 << mateQualities << endl;
}
else {
// end 2
fq2 << "@" << bam1.Name << "/2" <<endl;
fq2 << seq1 << endl;
fq2 << "+" << endl;
fq2 << qual1 << endl;
// end 1
fq1 << "@" << bam1.Name << "/1" <<endl;
fq1 << mateSequence << endl;
fq1 << "+" << endl;
fq1 << mateQualities << endl;
}
}
}
reader.Close();
}
void BamToFastq::PairedFastq() {
// open the 1st fastq file for writing
ofstream fq1(_fastq1.c_str(), ios::out);
if ( !fq1 ) {
cerr << "Error: The first fastq file (" << _fastq1 << ") could not be opened. Exiting!" << endl;
exit (1);
}
// open the 2nd fastq file for writing
ofstream fq2(_fastq2.c_str(), ios::out);
if ( !fq2 ) {
cerr << "Error: The second fastq file (" << _fastq2 << ") could not be opened. Exiting!" << endl;
exit (1);
}
// open the BAM file
BamReader reader;
reader.Open(_bamFile);
// rip through the BAM file and convert each mapped entry to BEDPE
BamAlignment bam1, bam2;
bool shouldConsumeReads = true;
while (true) {
if (shouldConsumeReads) {
if (!reader.GetNextAlignment(bam1) || !reader.GetNextAlignment(bam2)) break;
} else {
shouldConsumeReads = true;
}
if (bam1.Name != bam2.Name) {
while (bam1.Name != bam2.Name)
{
if (bam1.IsPaired())
{
cerr << "*****WARNING: Query " << bam1.Name
<< " is marked as paired, but its mate does not occur"
<< " next to it in your BAM file. Skipping. " << endl;
}
bam1 = bam2;
if (!reader.GetNextAlignment(bam2)) break;
shouldConsumeReads = false;
}
}
else if (bam1.IsPaired() && bam2.IsPaired()) {
// extract the sequence and qualities for the BAM "query"
string seq1 = bam1.QueryBases;
string qual1 = bam1.Qualities;
string seq2 = bam2.QueryBases;
string qual2 = bam2.Qualities;
if (bam1.IsReverseStrand() == true) {
reverseComplement(seq1);
reverseSequence(qual1);
}
if (bam2.IsReverseStrand() == true) {
reverseComplement(seq2);
reverseSequence(qual2);
}
fq1 << "@" << bam1.Name << "/1" << endl;
fq1 << seq1 << endl;
fq1 << "+" << endl;
fq1 << qual1 << endl;
fq2 << "@" << bam2.Name << "/2" << endl;
fq2 << seq2 << endl;
fq2 << "+" << endl;
fq2 << qual2 << endl;
}
}
reader.Close();
}
int main (int argc, char *argv[]) {
bool mapped =false;
bool unmapped=false;
const string usage=string(string(argv[0])+" [options] input.bam out.bam"+"\n\n"+
"This program takes a BAM file as input and produces\n"+
"another where the putative deaminated bases have\n"+
"have been cut\n"+
"\n"+
"Options:\n");
// "\t"+"-u , --unmapped" +"\n\t\t"+"For an unmapped bam file"+"\n"+
// "\t"+"-m , --mapped" +"\n\t\t"+"For an mapped bam file"+"\n");
if( (argc== 1) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cout<<"Usage:"<<endl;
cout<<usage<<endl;
cout<<""<<endl;
return 1;
}
// for(int i=1;i<(argc-1);i++){ //all but the last arg
// if(strcmp(argv[i],"-m") == 0 || strcmp(argv[i],"--mapped") == 0 ){
// mapped=true;
// continue;
// }
// if(strcmp(argv[i],"-u") == 0 || strcmp(argv[i],"--unmapped") == 0 ){
// unmapped=true;
// continue;
// }
// cerr<<"Unknown option "<<argv[i] <<" exiting"<<endl;
// return 1;
// }
if(argc != 3){
cerr<<"Error: Must specify the input and output BAM files";
return 1;
}
string inbamFile =argv[argc-2];
string outbamFile=argv[argc-1];
// if(!mapped && !unmapped){
// cerr << "Please specify whether you reads are mapped or unmapped" << endl;
// return 1;
// }
// if(mapped && unmapped){
// cerr << "Please specify either mapped or unmapped but not both" << endl;
// return 1;
// }
BamReader reader;
if ( !reader.Open(inbamFile) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
vector<RefData> testRefData=reader.GetReferenceData();
const SamHeader header = reader.GetHeader();
const RefVector references = reader.GetReferenceData();
BamWriter writer;
if ( !writer.Open(outbamFile, header, references) ) {
cerr << "Could not open output BAM file" << endl;
return 1;
}
BamAlignment al;
// BamAlignment al2;
// bool al2Null=true;
while ( reader.GetNextAlignment(al) ) {
if(al.IsPaired() ){
if(al.IsFirstMate() ){ //5' end, need to check first base only
if(al.IsReverseStrand()){ //
if(!al.IsMapped()){
cerr << "Cannot have reverse complemented unmapped reads: " <<al.Name<< endl;
//return 1;
}
int indexToCheck;
//last
indexToCheck=al.QueryBases.length()-1;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
}
}else{
int indexToCheck;
//first base
indexToCheck=0;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
}
}
}else{ //3' end, need to check last two bases only
if( al.IsSecondMate() ){
if(al.IsReverseStrand()){ //
if(!al.IsMapped()){
cerr << "Cannot have reverse complemented unmapped reads: " <<al.Name<< endl;
//return 1;
}
int indexToCheck;
//second to last
indexToCheck=al.QueryBases.length()-2;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
}else{
//last
indexToCheck=al.QueryBases.length()-1;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
}
}
}else{
int indexToCheck;
//second base
indexToCheck=1;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
}else{
//first base
indexToCheck=0;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
//al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
}
}
}
}else{
cerr << "Wrong state" << endl;
return 1;
}
}
}//end of paired end
else{//we consider single reads to have been sequenced from 5' to 3'
if(al.IsReverseStrand()){ //need to consider
if(!al.IsMapped()){
cerr << "Cannot have reverse complemented unmapped reads: " <<al.Name<< endl;
//return 1;
}
int indexToCheck;
//second base
indexToCheck=1;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"51 "<<al.QueryBases<<endl;
// cout<<"51 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
// cout<<"52 "<<al.QueryBases<<endl;
// cout<<"52 "<<al.Qualities<<endl;
}else{
//first base
indexToCheck=0;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"61 "<<al.QueryBases<<endl;
// cout<<"61 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
// cout<<"62 "<<al.QueryBases<<endl;
// cout<<"62 "<<al.Qualities<<endl;
}
}
//last
indexToCheck=al.QueryBases.length()-1;
if(toupper(al.QueryBases[indexToCheck]) == 'A'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"21 "<<al.QueryBases<<endl;
// cout<<"21 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
// cout<<"22 "<<al.QueryBases<<endl;
// cout<<"22 "<<al.Qualities<<endl;
}
}else{
int indexToCheck;
//first base
indexToCheck=0;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"11 "<<al.QueryBases<<endl;
// cout<<"11 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(indexToCheck+1);
al.Qualities = al.Qualities.substr(indexToCheck+1);
// cout<<"12 "<<al.QueryBases<<endl;
// cout<<"12 "<<al.Qualities<<endl;
}
//second to last
indexToCheck=al.QueryBases.length()-2;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"31 "<<al.QueryBases<<endl;
// cout<<"31 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
// cout<<"32 "<<al.QueryBases<<endl;
// cout<<"32 "<<al.Qualities<<endl;
}else{
//last
indexToCheck=al.QueryBases.length()-1;
if(toupper(al.QueryBases[indexToCheck]) == 'T'){
// al.Qualities[indexToCheck]=char(offset+baseQualForDeam);
// cout<<"41 "<<al.QueryBases<<endl;
// cout<<"41 "<<al.Qualities<<endl;
al.QueryBases = al.QueryBases.substr(0,indexToCheck);
al.Qualities = al.Qualities.substr(0, indexToCheck);
// cout<<"42 "<<al.QueryBases<<endl;
// cout<<"42 "<<al.Qualities<<endl;
}
}
}
}//end of single end
writer.SaveAlignment(al);
}// while ( reader.GetNextAlignment(al) ) {
reader.Close();
writer.Close();
return 0;
}
int main (int argc, char *argv[]) {
int minBaseQuality = 0;
string usage=string(""+string(argv[0])+" [in BAM file] [in VCF file] [chr name] [deam out BAM] [not deam out BAM]"+
"\nThis program divides aligned single end reads into potentially deaminated\n"+
"\nreads and the puts the rest into another bam file if the deaminated positions are not called as the alternative base in the VCF.\n"+
"\nThis is like filterDeaminatedVCF but it loads the VCF before then labels the reads instead of doing it on the fly\n"+
"\nwhich is good if you have many reads in the bam file.\n"+
"\nTip: if you do not need one of them, use /dev/null as your output\n"+
"\narguments:\n"+
"\t"+"--bq [base qual] : Minimum base quality to flag a deaminated site (Default: "+stringify(minBaseQuality)+")\n"+
"\t"+"--1000g [vcf file] : VCF file from 1000g to get the putative A and T positions in modern humans (Default: "+vcf1000g+")\n"+
"\n");
if(argc == 1 ||
argc < 4 ||
(argc == 2 && (string(argv[0]) == "-h" || string(argv[0]) == "--help") )
){
cerr << "Usage "<<usage<<endl;
return 1;
}
for(int i=1;i<(argc-2);i++){
if(string(argv[i]) == "--bq"){
minBaseQuality=destringify<int>(argv[i+1]);
i++;
continue;
}
if(string(argv[i]) == "--1000g"){
vcf1000g=string(argv[i+1]);
i++;
continue;
}
}
unsigned int maxSizeChromosome=250000000;//larger than chr1 hg19
bool * hasCnoT;
bool * hasGnoA;
bool * thousandGenomesHasA;
bool * thousandGenomesHasT;
cerr<<"Trying to allocating memory"<<endl;
try{
hasCnoT = new bool[ maxSizeChromosome ];
hasGnoA = new bool[ maxSizeChromosome ];
thousandGenomesHasA = new bool[ maxSizeChromosome ];
thousandGenomesHasT = new bool[ maxSizeChromosome ];
}catch(bad_alloc& exc){
cerr<<"ERROR: allocating memory failed"<<endl;
return 1;
}
cerr<<"Success in allocating memory"<<endl;
for(unsigned int i = 0;i<maxSizeChromosome;i++){
hasCnoT[i]=false;
hasGnoA[i]=false;
thousandGenomesHasA[i]=false;
thousandGenomesHasT[i]=false;
}
string bamfiletopen = string( argv[ argc-5 ] );
string vcffiletopen = string( argv[ argc-4 ] );
string chrname = string( argv[ argc-3 ] );
string deambam = string( argv[ argc-2 ] );
string nondeambam = string( argv[ argc-1 ] );
cerr<<"Reading consensus VCF "<<vcffiletopen<<" ... "<<endl;
VCFreader vcfr (vcffiletopen,
// vcffiletopen+".tbi",
// chrname,
// 1,
// maxSizeChromosome,
0);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
if(toprint->getRef().length() != 1 )
continue;
//if the VCF has a at least one G but no A
if( toprint->hasAtLeastOneG() &&
!toprint->hasAtLeastOneA() ){
hasGnoA[ toprint->getPosition() ] =true;
}
if( toprint->hasAtLeastOneC() &&
!toprint->hasAtLeastOneT() ){
hasCnoT[ toprint->getPosition() ] =true;
}
}
cerr<<"done reading VCF"<<endl;
cerr<<"Reading 1000g VCF :"<<vcf1000g<<" ..."<<endl;
string line1000g;
ifstream myFile1000g;
myFile1000g.open(vcf1000g.c_str(), ios::in);
if (myFile1000g.is_open()){
while ( getline (myFile1000g,line1000g)){
vector<string> fields=allTokens(line1000g,'\t');
//0 chr
//1 pos
//2 id
//3 ref
//4 alt
//check if same chr
if(fields[0] != chrname){
cerr <<"Error, wrong chromosome in 1000g file for line= "<<line1000g<<endl;
return 1;
}
//skip indels
if(fields[3].size() != 1 ||
fields[4].size() != 1 )
continue;
char ref=toupper(fields[3][0]);
char alt=toupper(fields[4][0]);
unsigned int pos=destringify<unsigned int>( fields[1] );
thousandGenomesHasA[ pos ] = ( (ref=='A') || (alt=='A') );
thousandGenomesHasT[ pos ] = ( (ref=='T') || (alt=='T') );
}
myFile1000g.close();
}else{
cerr <<"Unable to open file "<<vcf1000g<<endl;
return 1;
}
cerr<<"done reading 1000g VCF"<<endl;
BamReader reader;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM file"<< bamfiletopen << endl;
return 1;
}
//positioning the bam file
int refid=reader.GetReferenceID(chrname);
if(refid < 0){
cerr << "Cannot retrieve the reference ID for "<< chrname << endl;
return 1;
}
//cout<<"redif "<<refid<<endl;
//setting the BAM reader at that position
reader.SetRegion(refid,
0,
refid,
-1);
vector<RefData> testRefData=reader.GetReferenceData();
const SamHeader header = reader.GetHeader();
const RefVector references = reader.GetReferenceData();
BamWriter writerDeam;
if ( !writerDeam.Open(deambam, header, references) ) {
cerr << "Could not open output BAM file" << endl;
return 1;
}
BamWriter writerNoDeam;
if ( !writerNoDeam.Open(nondeambam, header, references) ) {
cerr << "Could not open output BAM file" << endl;
return 1;
}
unsigned int totalReads =0;
unsigned int deaminatedReads =0;
unsigned int ndeaminatedReads =0;
unsigned int skipped =0;
//iterating over the alignments for these regions
BamAlignment al;
int i;
while ( reader.GetNextAlignment(al) ) {
// cerr<<al.Name<<endl;
//skip unmapped
if(!al.IsMapped()){
skipped++;
continue;
}
//skip paired end !
if(al.IsPaired() ){
continue;
// cerr<<"Paired end not yet coded"<<endl;
// return 1;
}
string reconstructedReference = reconstructRef(&al);
char refeBase;
char readBase;
bool isDeaminated;
if(al.Qualities.size() != reconstructedReference.size()){
cerr<<"Quality line is not the same size as the reconstructed reference"<<endl;
return 1;
}
isDeaminated=false;
if(al.IsReverseStrand()){
//first base next to 3'
i = 0 ;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
if( hasGnoA[al.Position+1] &&
!thousandGenomesHasA[al.Position+1] )
isDeaminated=true;
// transformRef(&refeBase,&readBase);
// vcfr.repositionIterator(chrname,al.Position+1,al.Position+1);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// // cout<<*toprint<<endl;
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<"Problem1 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// //if the VCF has a at least one G but no A
// if( toprint->hasAtLeastOneG() &&
// !toprint->hasAtLeastOneA() ){
// isDeaminated=true;
// }
// }
}
//second base next to 3'
i = 1;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'G' &&
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
if( hasGnoA[al.Position+2] &&
!thousandGenomesHasA[al.Position+2] )
isDeaminated=true;
// transformRef(&refeBase,&readBase);
// vcfr.repositionIterator(chrname,al.Position+2,al.Position+2);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// // cout<<*toprint<<endl;
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<"Problem2 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// //if the VCF has at least one G but no A
// // if(toprint->hasAtLeastOneG() &&
// // toprint->getAlt().find("A") == string::npos){
// if( toprint->hasAtLeastOneG() &&
// !toprint->hasAtLeastOneA() ){
// isDeaminated=true;
// }
// }
}
//last base next to 5'
i = (al.QueryBases.length()-1) ;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'G' &&
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,0);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
if(hasGnoA[positionJump] &&
!thousandGenomesHasA[positionJump] )
isDeaminated=true;
// vcfr.repositionIterator(chrname,positionJump,positionJump);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<lengthMatches<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<positionJump<<endl;
// cerr<<"Problem3 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// //if the VCF has at least one G but no A
// if( toprint->hasAtLeastOneG() &&
// !toprint->hasAtLeastOneA() ){
// isDeaminated=true;
// }
// }
}
}else{
//first base next to 5'
i = 0;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'C'
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
// transformRef(&refeBase,&readBase);
if(hasCnoT[al.Position+1] &&
!thousandGenomesHasT[al.Position+1] )
isDeaminated=true;
// vcfr.repositionIterator(chrname,al.Position+1,al.Position+1);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// //cout<<*toprint<<endl;
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<"Problem4 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// //if the VCF has at least one C but no T
// if( toprint->hasAtLeastOneC() &&
// !toprint->hasAtLeastOneT() ){
// isDeaminated=true;
// }
// }
//cout<<al.Position+
}
//second last base next to 3'
i = (al.QueryBases.length()-2);
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'C' &&
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
//transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,1);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
if(hasCnoT[positionJump] &&
!thousandGenomesHasT[positionJump] )
isDeaminated=true;
// vcfr.repositionIterator(chrname,positionJump,positionJump);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<lengthMatches<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<positionJump<<endl;
// cerr<<"Problem5 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// if( toprint->hasAtLeastOneC() &&
// !toprint->hasAtLeastOneT() ){
// isDeaminated=true;
// }
// }
}
//last base next to 3'
i = (al.QueryBases.length()-1);
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//&& refeBase == 'C'
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,0);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
if(hasCnoT[positionJump] &&
!thousandGenomesHasT[positionJump] )
isDeaminated=true;
// vcfr.repositionIterator(chrname,positionJump,positionJump);
// while(vcfr.hasData()){
// SimpleVCF * toprint=vcfr.getData();
// //skip deletions in the alt
// if(toprint->getRef().length() != 1 )
// continue;
// if(toprint->getRef()[0] != refeBase){
// cerr<<reconstructedReference<<endl;
// cerr<<al.Position<<endl;
// cerr<<lengthMatches<<endl;
// cerr<<numberOfDeletions(&al)<<endl;
// cerr<<positionJump<<endl;
// cerr<<"Problem6 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
// exit(1);
// }
// if( toprint->hasAtLeastOneC() &&
// !toprint->hasAtLeastOneT() ){
// isDeaminated=true;
// }
// }
}
}
totalReads++;
if(isDeaminated){
deaminatedReads++;
writerDeam.SaveAlignment(al);
}else{
ndeaminatedReads++;
writerNoDeam.SaveAlignment(al);
}
}//end for each read
reader.Close();
writerDeam.Close();
writerNoDeam.Close();
delete(hasCnoT);
delete(hasGnoA);
cerr<<"Program finished sucessfully, out of "<<totalReads<<" mapped reads (skipped: "<<skipped<<" reads) we flagged "<<deaminatedReads<<" as deaminated and "<<ndeaminatedReads<<" as not deaminated"<<endl;
return 0;
}
// close requested BAM files
bool BamMultiReaderPrivate::CloseFiles(const std::vector<std::string>& filenames)
{
bool errorsEncountered = false;
m_errorString.clear();
// iterate over filenames
std::vector<std::string>::const_iterator filesIter = filenames.begin();
std::vector<std::string>::const_iterator filesEnd = filenames.end();
for (; filesIter != filesEnd; ++filesIter) {
const std::string& filename = (*filesIter);
if (filename.empty()) continue;
// iterate over readers
std::vector<MergeItem>::iterator readerIter = m_readers.begin();
std::vector<MergeItem>::iterator readerEnd = m_readers.end();
for (; readerIter != readerEnd; ++readerIter) {
MergeItem& item = (*readerIter);
BamReader* reader = item.Reader;
if (reader == 0) continue;
// if reader matches requested filename
if (reader->GetFilename() == filename) {
// remove reader's entry from alignment cache
m_alignmentCache->Remove(reader);
// clean up reader & its alignment
if (!reader->Close()) {
m_errorString.append(1, '\t');
m_errorString.append(reader->GetErrorString());
m_errorString.append(1, '\n');
errorsEncountered = true;
}
delete reader;
reader = 0;
// delete reader's alignment entry
BamAlignment* alignment = item.Alignment;
delete alignment;
alignment = 0;
// remove reader from reader list
m_readers.erase(readerIter);
// on match, just go on to next filename
// (no need to keep looking and item iterator is invalid now anyway)
break;
}
}
}
// make sure we clean up properly if all readers were closed
if (m_readers.empty()) {
// clean up merger
if (m_alignmentCache) {
m_alignmentCache->Clear();
delete m_alignmentCache;
m_alignmentCache = 0;
}
// reset merge flags
m_hasUserMergeOrder = false;
m_mergeOrder = BamMultiReader::RoundRobinMerge;
}
// return whether all readers closed OK
return !errorsEncountered;
}
//{{{ bool merge_sorted_files(string out_file_name,
bool merge_sorted_files(string out_file_name,
int buff_count,
string header_text,
RefVector &ref)
{
map<string,BamReader*> bam_readers;
priority_queue< BamAlignment, vector<BamAlignment>, inter_chrom_rev_sort > q;
for (int i = 0; i < buff_count; ++i) {
stringstream temp_name;
temp_name << out_file_name << i;
BamReader *reader = new BamReader();
if ( !reader->Open(temp_name.str()) ) {
cerr << "sort ERROR: could not open " <<
temp_name.str() << " for reading... Aborting." << endl;
return false;
}
bam_readers[temp_name.str()] = reader;
// place an item from each bam onto the q
BamAlignment al;
if (reader->GetNextAlignment(al))
q.push(al);
}
BamWriter merged_writer;
if ( !merged_writer.Open(out_file_name, header_text, ref) ) {
cerr << "sort ERROR: could not open " << out_file_name
<< " for writing." << endl;
return false;
}
while (!q.empty()) {
BamAlignment al = q.top();
q.pop();
merged_writer.SaveAlignment(al);
BamReader *reader = bam_readers[al.Filename];
BamAlignment new_al;
if (reader->GetNextAlignment(new_al))
q.push(new_al);
}
merged_writer.Close();
//close and remove temp files
map<string,BamReader*>::iterator it;
for (it = bam_readers.begin(); it != bam_readers.end(); ++it) {
BamReader *reader = it->second;
reader->Close();
delete reader;
remove(it->first.c_str());
}
return true;
}
void BedIntersect::IntersectBam(string bamFile) {
// load the "B" bed file into a map so
// that we can easily compare "A" to it for overlaps
_bedB = new BedFile(_bedBFile);
_bedB->loadBedFileIntoMap();
// create a dummy BED A file for printing purposes if not
// using BAM output.
if (_bamOutput == false) {
_bedA = new BedFile(_bedAFile);
_bedA->bedType = 12;
}
// open the BAM file
BamReader reader;
BamWriter writer;
reader.Open(bamFile);
// get header & reference information
string bamHeader = reader.GetHeaderText();
RefVector refs = reader.GetReferenceData();
// open a BAM output to stdout if we are writing BAM
if (_bamOutput == true) {
// set compression mode
BamWriter::CompressionMode compressionMode = BamWriter::Compressed;
if ( _isUncompressedBam ) compressionMode = BamWriter::Uncompressed;
writer.SetCompressionMode(compressionMode);
// open our BAM writer
writer.Open("stdout", bamHeader, refs);
}
vector<BED> hits;
// reserve some space
hits.reserve(100);
BamAlignment bam;
// get each set of alignments for each pair.
while (reader.GetNextAlignment(bam)) {
// save an unaligned read if -v
if (!bam.IsMapped()) {
if (_noHit == true)
writer.SaveAlignment(bam);
continue;
}
// break alignment into discrete blocks,
bedVector bed_blocks;
string chrom = refs.at(bam.RefID).RefName;
GetBamBlocks(bam, chrom, bed_blocks, false, true);
// create a basic BED entry from the BAM alignment
BED bed;
MakeBedFromBam(bam, chrom, bed_blocks, bed);
bool overlapsFound = false;
if ((_bamOutput == true) && (_obeySplits == false))
{
overlapsFound = _bedB->anyHits(bed.chrom, bed.start, bed.end,
bed.strand, _sameStrand, _diffStrand,
_overlapFraction, _reciprocal);
}
else if ( ((_bamOutput == true) && (_obeySplits == true)) ||
((_bamOutput == false) && (_obeySplits == true)) )
{
// find the hits that overlap with the full span of the blocked BED
_bedB->allHits(bed.chrom, bed.start, bed.end, bed.strand,
hits, _sameStrand, _diffStrand,
_overlapFraction, _reciprocal);
// find the overlaps between the block in A and B
overlapsFound = FindBlockedOverlaps(bed, bed_blocks, hits, _bamOutput);
}
else if ((_bamOutput == false) && (_obeySplits == false))
{
FindOverlaps(bed, hits);
}
// save the BAM alignment if overlap reqs. were met
if (_bamOutput == true) {
if ((overlapsFound == true) && (_noHit == false))
writer.SaveAlignment(bam);
else if ((overlapsFound == false) && (_noHit == true))
writer.SaveAlignment(bam);
}
hits.clear();
}
// close the relevant BAM files.
reader.Close();
if (_bamOutput == true) {
writer.Close();
}
}
int main (int argc, char *argv[]) {
int minBaseQuality = 0;
string usage=string(""+string(argv[0])+" [in BAM file] [in VCF file] [chr name] [deam out BAM] [not deam out BAM]"+
"\nThis program divides aligned single end reads into potentially deaminated\n"+
"\nreads and the puts the rest into another bam file if the deaminated positions are not called as the alternative base in the VCF.\n"+
"\nTip: if you do not need one of them, use /dev/null as your output\n"+
"arguments:\n"+
"\t"+"--bq [base qual] : Minimum base quality to flag a deaminated site (Default: "+stringify(minBaseQuality)+")\n"+
"\n");
if(argc == 1 ||
argc < 4 ||
(argc == 2 && (string(argv[0]) == "-h" || string(argv[0]) == "--help") )
){
cerr << "Usage "<<usage<<endl;
return 1;
}
for(int i=1;i<(argc-2);i++){
if(string(argv[i]) == "--bq"){
minBaseQuality=destringify<int>(argv[i+1]);
i++;
continue;
}
}
string bamfiletopen = string( argv[ argc-5 ] );
string vcffiletopen = string( argv[ argc-4 ] );
string chrname = string( argv[ argc-3 ] );
string deambam = string( argv[ argc-2 ] );
string nondeambam = string( argv[ argc-1 ] );
//dummy reader, will need to reposition anyway
VCFreader vcfr (vcffiletopen,
vcffiletopen+".tbi",
chrname,
1,
1,
0);
BamReader reader;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM file"<< bamfiletopen << endl;
return 1;
}
// if ( !reader.LocateIndex() ) {
// cerr << "The index for the BAM file cannot be located" << endl;
// return 1;
// }
// if ( !reader.HasIndex() ) {
// cerr << "The BAM file has not been indexed." << endl;
// return 1;
// }
//positioning the bam file
int refid=reader.GetReferenceID(chrname);
if(refid < 0){
cerr << "Cannot retrieve the reference ID for "<< chrname << endl;
return 1;
}
//cout<<"redif "<<refid<<endl;
//setting the BAM reader at that position
reader.SetRegion(refid,
0,
refid,
-1);
vector<RefData> testRefData=reader.GetReferenceData();
const SamHeader header = reader.GetHeader();
const RefVector references = reader.GetReferenceData();
BamWriter writerDeam;
if ( !writerDeam.Open(deambam, header, references) ) {
cerr << "Could not open output BAM file" << endl;
return 1;
}
BamWriter writerNoDeam;
if ( !writerNoDeam.Open(nondeambam, header, references) ) {
cerr << "Could not open output BAM file" << endl;
return 1;
}
unsigned int totalReads =0;
unsigned int deaminatedReads =0;
unsigned int ndeaminatedReads =0;
unsigned int skipped =0;
//iterating over the alignments for these regions
BamAlignment al;
int i;
while ( reader.GetNextAlignment(al) ) {
// cerr<<al.Name<<endl;
//skip unmapped
if(!al.IsMapped()){
skipped++;
continue;
}
//skip paired end !
if(al.IsPaired() ){
continue;
// cerr<<"Paired end not yet coded"<<endl;
// return 1;
}
string reconstructedReference = reconstructRef(&al);
char refeBase;
char readBase;
bool isDeaminated;
if(al.Qualities.size() != reconstructedReference.size()){
cerr<<"Quality line is not the same size as the reconstructed reference"<<endl;
return 1;
}
isDeaminated=false;
if(al.IsReverseStrand()){
//first base next to 3'
i = 0 ;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
vcfr.repositionIterator(chrname,al.Position+1,al.Position+1);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
// cout<<*toprint<<endl;
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<"Problem1 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
//if the VCF has a at least one G but no A
if( toprint->hasAtLeastOneG() &&
!toprint->hasAtLeastOneA() ){
isDeaminated=true;
}
}
}
//second base next to 3'
i = 1;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'G' &&
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
vcfr.repositionIterator(chrname,al.Position+2,al.Position+2);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
// cout<<*toprint<<endl;
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<"Problem2 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
//if the VCF has at least one G but no A
// if(toprint->hasAtLeastOneG() &&
// toprint->getAlt().find("A") == string::npos){
if( toprint->hasAtLeastOneG() &&
!toprint->hasAtLeastOneA() ){
isDeaminated=true;
}
}
}
//last base next to 5'
i = (al.QueryBases.length()-1) ;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'G' &&
if( readBase == 'A' && int(al.Qualities[i]-offset) >= minBaseQuality){ //isDeaminated=true; }
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,0);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
vcfr.repositionIterator(chrname,positionJump,positionJump);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<lengthMatches<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<positionJump<<endl;
cerr<<"Problem3 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
//if the VCF has at least one G but no A
if( toprint->hasAtLeastOneG() &&
!toprint->hasAtLeastOneA() ){
isDeaminated=true;
}
}
}
}else{
//first base next to 5'
i = 0;
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'C'
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
vcfr.repositionIterator(chrname,al.Position+1,al.Position+1);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
//cout<<*toprint<<endl;
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<"Problem4 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
//if the VCF has at least one C but no T
if( toprint->hasAtLeastOneC() &&
!toprint->hasAtLeastOneT() ){
isDeaminated=true;
}
}
//cout<<al.Position+
}
//second last base next to 3'
i = (al.QueryBases.length()-2);
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//refeBase == 'C' &&
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,1);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
vcfr.repositionIterator(chrname,positionJump,positionJump);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<lengthMatches<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<positionJump<<endl;
cerr<<"Problem5 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
if( toprint->hasAtLeastOneC() &&
!toprint->hasAtLeastOneT() ){
isDeaminated=true;
}
}
}
//last base next to 3'
i = (al.QueryBases.length()-1);
refeBase=toupper(reconstructedReference[i]);
readBase=toupper( al.QueryBases[i]);
//&& refeBase == 'C'
if( readBase == 'T' && int(al.Qualities[i]-offset) >= minBaseQuality){
if( skipAlign(reconstructedReference,&al,&skipped) ){ continue; }
transformRef(&refeBase,&readBase);
int lengthMatches=countMatchesRecons(reconstructedReference,0);
int positionJump = al.Position+lengthMatches+numberOfDeletions(&al);
vcfr.repositionIterator(chrname,positionJump,positionJump);
while(vcfr.hasData()){
SimpleVCF * toprint=vcfr.getData();
//skip deletions in the alt
if(toprint->getRef().length() != 1 )
continue;
if(toprint->getRef()[0] != refeBase){
cerr<<reconstructedReference<<endl;
cerr<<al.Position<<endl;
cerr<<lengthMatches<<endl;
cerr<<numberOfDeletions(&al)<<endl;
cerr<<positionJump<<endl;
cerr<<"Problem6 position "<<*toprint<<" does not have a "<<refeBase<<" as reference allele for read "<<al.Name<<endl;
exit(1);
}
if( toprint->hasAtLeastOneC() &&
!toprint->hasAtLeastOneT() ){
isDeaminated=true;
}
}
}
}
totalReads++;
if(isDeaminated){
deaminatedReads++;
writerDeam.SaveAlignment(al);
}else{
ndeaminatedReads++;
writerNoDeam.SaveAlignment(al);
}
}//end for each read
reader.Close();
writerDeam.Close();
writerNoDeam.Close();
cerr<<"Program finished sucessfully, out of "<<totalReads<<" mapped reads (skipped: "<<skipped<<" reads) we flagged "<<deaminatedReads<<" as deaminated and "<<ndeaminatedReads<<" as not deaminated"<<endl;
return 0;
}
// close the BAM files
void BamMultiReader::Close(void) {
for (vector<pair<BamReader*, BamAlignment*> >::iterator it = readers.begin(); it != readers.end(); ++it) {
BamReader* reader = it->first;
reader->Close(); // close the reader
}
}
int main (int argc, char *argv[]) {
if( (argc!= 3) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cerr<<"Usage:splitByRG [in bam] [out prefix]"<<endl<<"this program creates one bam file per RG in the with the outprefix\nFor example splitByRG in.bam out will create\nout.rg1.bam\nout.rg2.bam\n"<<endl;
return 1;
}
string bamfiletopen = string(argv[1]);
// if(!strEndsWith(bamfiletopen,".bam")){
// }
string bamDirOutPrefix = string(argv[2]);
map<string,BamWriter *> rg2BamWriter;
// if(!isDirectory(bamDirOut)){
// cerr<<"ERROR: the out directory does not exist"<<endl;
// return 1;
// }
BamReader reader;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
SamHeader header = reader.GetHeader();
const RefVector references = reader.GetReferenceData();
vector<RefData> refData=reader.GetReferenceData();
string pID = "splitByRG";
string pName = "splitByRG";
string pCommandLine = "";
for(int i=0;i<(argc);i++){
pCommandLine += (string(argv[i])+" ");
}
putProgramInHeader(&header,pID,pName,pCommandLine,returnGitHubVersion(string(argv[0]),".."));
SamReadGroupDictionary srgd=header.ReadGroups;
for(SamReadGroupConstIterator srgci=srgd.ConstBegin();
srgci<srgd.ConstEnd();
srgci++){
//cout<<*srgci<<endl;
const SamReadGroup rg = (*srgci);
//cout<<rg.ID<<endl;
rg2BamWriter[rg.ID] = new BamWriter();
rg2BamWriter[rg.ID]->Open(bamDirOutPrefix+"."+rg.ID+".bam",header,references);
}
BamAlignment al;
unsigned int total=0;
while ( reader.GetNextAlignment(al) ) {
// al.SetIsFailedQC(false);
// writer.SaveAlignment(al);
// if(al.IsMapped () ){
// if(rg2BamWriter.find(refData[al.RefID].RefName) == rg2BamWriter.end()){ //new
// rg2BamWriter[refData[al.RefID].RefName] = new BamWriter();
// if ( !rg2BamWriter[refData[al.RefID].RefName]->Open(bamDirOutPrefix+"."+refData[al.RefID].RefName+".bam",header,references) ) {
// cerr << "Could not open output BAM file "<< bamDirOutPrefix<<"."<<refData[al.RefID].RefName<<".bam" << endl;
// return 1;
// }
// }else{
// rg2BamWriter[refData[al.RefID].RefName]->SaveAlignment(al);
// }
// }else{
// unmapped.SaveAlignment(al);
// }
if(al.HasTag("RG")){
string rgTag;
al.GetTag("RG",rgTag);
//cout<<rgTag<<endl;
if(rg2BamWriter.find(rgTag) == rg2BamWriter.end()){ //new
cerr<<"Found new RG "<<rgTag<<endl;
rg2BamWriter[rgTag] = new BamWriter();
if ( !rg2BamWriter[rgTag]->Open(bamDirOutPrefix+"."+rgTag+".bam",header,references) ) {
cerr << "Could not open output BAM file "<< bamDirOutPrefix<<"."<<rgTag<<".bam" << endl;
return 1;
}
rg2BamWriter[rgTag]->SaveAlignment(al);
}else{
rg2BamWriter[rgTag]->SaveAlignment(al);
}
}else{
string rgTag="unknown";
//cout<<rgTag<<endl;
if(rg2BamWriter.find(rgTag) == rg2BamWriter.end()){ //new
cerr<<"Found new RG "<<rgTag<<endl;
rg2BamWriter[rgTag] = new BamWriter();
if ( !rg2BamWriter[rgTag]->Open(bamDirOutPrefix+"."+rgTag+".bam",header,references) ) {
cerr << "Could not open output BAM file "<< bamDirOutPrefix<<"."<<rgTag<<".bam" << endl;
return 1;
}
rg2BamWriter[rgTag]->SaveAlignment(al);
}else{
rg2BamWriter[rgTag]->SaveAlignment(al);
}
// cerr << "Cannot get RG tag for " << al.Name<<endl;
// return 1;
}
total++;
} //while al
reader.Close();
// writer.Close();
// unmapped.Close();
map<string,BamWriter *>::iterator rg2BamWriterIt;
for (rg2BamWriterIt =rg2BamWriter.begin();
rg2BamWriterIt!=rg2BamWriter.end();
rg2BamWriterIt++){
rg2BamWriterIt->second->Close();
}
cerr<<"Wrote succesfully "<<total<<" reads"<<endl;
return 0;
}
int main (int argc, char *argv[]) {
if( (argc== 1) ||
(argc== 2 && string(argv[1]) == "-h") ||
(argc== 2 && string(argv[1]) == "-help") ||
(argc== 2 && string(argv[1]) == "--help") ){
cout<<"Usage:"<<endl;
cout<<""<<endl;
cout<<"plotQualScore input.bam"<<endl;
return 1;
}
string bamfiletopen = string(argv[1]);
BamReader reader;
if ( !reader.Open(bamfiletopen) ) {
cerr << "Could not open input BAM files." << endl;
return 1;
}
// if ( !reader.LocateIndex() ){
// cerr << "warning: cannot locate index for file " << bamfiletopen<<endl;
// //return 1;
// }
BamAlignment al;
BamAlignment al2;
bool unsurePEorSE=true;
bool pe=true;
int strLength=-1;
int vecLengthToUse=-1;
map<short,unsigned long> ** counterA = 0;
map<short,unsigned long> ** counterC = 0;
map<short,unsigned long> ** counterG = 0;
map<short,unsigned long> ** counterT = 0;
int lengthIndex1=0;
int lengthIndex2=0;
string seqInd1;
string seqInd2;
string qualInd1;
string qualInd2;
int offsetInd2;
while ( reader.GetNextAlignment(al) ) {
if(unsurePEorSE){
strLength=al.QueryBases.length();
if(al.IsPaired()){
pe=true;
vecLengthToUse=2*strLength;
}else{
pe=false;
vecLengthToUse=strLength;
}
string index1;
string index2;
if(al.HasTag("XI")){
al.GetTag("XI",index1);
vecLengthToUse+=index1.length();
lengthIndex1=index1.length();
}
if(al.HasTag("XJ")){
al.GetTag("XJ",index2);
vecLengthToUse+=index2.length();
lengthIndex2=index2.length();
}
counterA = new map<short,unsigned long> * [vecLengthToUse];
counterC = new map<short,unsigned long> * [vecLengthToUse];
counterG = new map<short,unsigned long> * [vecLengthToUse];
counterT = new map<short,unsigned long> * [vecLengthToUse];
for(int i=0;i<vecLengthToUse;i++){
counterA[i]=new map<short,unsigned long> ();
counterC[i]=new map<short,unsigned long> ();
counterG[i]=new map<short,unsigned long> ();
counterT[i]=new map<short,unsigned long> ();
for(short k=minQualScore;k<=maxQualScore;k++){
(*counterA[i])[k]=0;
(*counterC[i])[k]=0;
(*counterG[i])[k]=0;
(*counterT[i])[k]=0;
}
}
unsurePEorSE=false;
}else{
if(pe &&
!al.IsPaired()){
cerr << "Cannot have unpaired reads in PE mode" << endl;
return 1;
}
if(!pe &&
al.IsPaired()){
cerr << "Cannot have unpaired reads in SE mode" << endl;
return 1;
}
}
if(al.QueryBases.length() != al.Qualities.length()){
cerr << "Cannot have different lengths for sequence and quality" << endl;
return 1;
}
if(int(al.QueryBases.length()) != strLength){
cerr << "Cannot have different lengths for sequence and quality" << endl;
return 1;
}
if(pe){
if(al.IsFirstMate()){
reader.GetNextAlignment(al2);
if(al2.QueryBases.length() != al2.Qualities.length()){
cerr << "Cannot have different lengths for sequence and quality" << endl;
return 1;
}
}else{
cerr << "First read should be the first mate" << endl;
return 1;
}
}
//cycle
for(unsigned int i=0;i<al.QueryBases.length();i++){
short x=(short(al.Qualities[i])-qualOffset);
if(al.QueryBases[i] == 'A'){
(*counterA[i])[x]++;
}
if(al.QueryBases[i] == 'C'){
(*counterC[i])[x]++;
}
if(al.QueryBases[i] == 'G'){
(*counterG[i])[x]++;
}
if(al.QueryBases[i] == 'T'){
(*counterT[i])[x]++;
}
}
//The indices for al and al2 should hopefully be the same
if(lengthIndex1>0){
al.GetTag("XI",seqInd1);
al.GetTag("YI",qualInd1);
int j;
for(int i=0;i<lengthIndex1;i++){
j=i+al.QueryBases.length();
short x=(short(qualInd1[i])-qualOffset);
if(seqInd1[i] == 'A'){
(*counterA[j])[x]++;
}
if(seqInd1[i] == 'C'){
(*counterC[j])[x]++;
}
if(seqInd1[i] == 'G'){
(*counterG[j])[x]++;
}
if(seqInd1[i] == 'T'){
(*counterT[j])[x]++;
}
}
}
if(pe){
offsetInd2=al.QueryBases.length()+lengthIndex1+al2.QueryBases.length();
int j;
for(unsigned int i=0;i<al2.QueryBases.length();i++){
j=i+al.QueryBases.length()+lengthIndex1;
short x=(short(al2.Qualities[i])-qualOffset);
if(al2.QueryBases[i] == 'A'){
(*counterA[j])[x]++;
}
if(al2.QueryBases[i] == 'C'){
(*counterC[j])[x]++;
}
if(al2.QueryBases[i] == 'G'){
(*counterG[j])[x]++;
}
if(al2.QueryBases[i] == 'T'){
(*counterT[j])[x]++;
}
}
}else{
offsetInd2=al.QueryBases.length()+lengthIndex1;
}
//The indices for al and al2 should hopefully be the same
if(lengthIndex2>0){
al.GetTag("XJ",seqInd2);
al.GetTag("YJ",qualInd2);
int j;
for(int i=0;i<lengthIndex2;i++){
j=offsetInd2+i;
short x=(short(qualInd2[i])-qualOffset);
if(seqInd2[i] == 'A'){
(*counterA[j])[x]++;
}
if(seqInd2[i] == 'C'){
(*counterC[j])[x]++;
}
if(seqInd2[i] == 'G'){
(*counterG[j])[x]++;
}
if(seqInd2[i] == 'T'){
(*counterT[j])[x]++;
}
}
}
}
reader.Close();
cout<<"cycle\t"<<"nuc\t";
for(short k=minQualScore;k<maxQualScore;k++){
cout<<k<<"\t";
}
cout<<maxQualScore<<endl;
for(int i=0;i<vecLengthToUse;i++){
cout<<(i+1)<<"\t";
cout<<"A\t";
for(short k=minQualScore;k<maxQualScore;k++){
cout<<(*counterA[i])[k]<<"\t";
}
cout<<(*counterA[i])[maxQualScore]<<endl;
cout<<(i+1)<<"\t";
cout<<"C\t";
for(short k=minQualScore;k<maxQualScore;k++){
cout<<(*counterC[i])[k]<<"\t";
}
cout<<(*counterC[i])[maxQualScore]<<endl;
cout<<(i+1)<<"\t";
cout<<"G\t";
for(short k=minQualScore;k<maxQualScore;k++){
cout<<(*counterG[i])[k]<<"\t";
}
cout<<(*counterG[i])[maxQualScore]<<endl;
cout<<(i+1)<<"\t";
cout<<"T\t";
for(short k=minQualScore;k<maxQualScore;k++){
cout<<(*counterT[i])[k]<<"\t";
}
cout<<(*counterT[i])[maxQualScore]<<endl;
}
return 0;
}
