/*
* The MIT License
*
* Copyright (c) 2011 The Broad Institute
*
* Permission is hereby granted, free of charge, to any person obtaining a copy
* of this software and associated documentation files (the "Software"), to deal
* in the Software without restriction, including without limitation the rights
* to use, copy, modify, merge, publish, distribute, sublicense, and/or sell
* copies of the Software, and to permit persons to whom the Software is
* furnished to do so, subject to the following conditions:
*
* The above copyright notice and this permission notice shall be included in
* all copies or substantial portions of the Software.
*
* THE SOFTWARE IS PROVIDED "AS IS", WITHOUT WARRANTY OF ANY KIND, EXPRESS OR
* IMPLIED, INCLUDING BUT NOT LIMITED TO THE WARRANTIES OF MERCHANTABILITY,
* FITNESS FOR A PARTICULAR PURPOSE AND NONINFRINGEMENT. IN NO EVENT SHALL THE
* AUTHORS OR COPYRIGHT HOLDERS BE LIABLE FOR ANY CLAIM, DAMAGES OR OTHER
* LIABILITY, WHETHER IN AN ACTION OF CONTRACT, TORT OR OTHERWISE, ARISING FROM,
* OUT OF OR IN CONNECTION WITH THE SOFTWARE OR THE USE OR OTHER DEALINGS IN
* THE SOFTWARE.
*/
package picard.sam;
import htsjdk.samtools.Cigar;
import htsjdk.samtools.CigarElement;
import htsjdk.samtools.CigarOperator;
import htsjdk.samtools.SAMRecord;
import htsjdk.samtools.SAMRecordQueryNameComparator;
import htsjdk.samtools.SAMTag;
import htsjdk.samtools.SAMUtils;
import htsjdk.samtools.filter.FilteringIterator;
import htsjdk.samtools.filter.SamRecordFilter;
import htsjdk.samtools.util.CloseableIterator;
import htsjdk.samtools.util.PeekableIterator;
import picard.PicardException;
import java.util.ArrayList;
import java.util.Arrays;
import java.util.List;
import java.util.NoSuchElementException;
/**
* Iterate over queryname-sorted SAM, and return each group of reads with the same queryname. Unmapped reads
* are filtered out, as are alignments that don't seem to match any part of the reference.
* If there are multiple hits for the same read, and the first and second ends need to be correlated,
* then they are sorted by hit index. Supplemental alignments are discarded, with a logged message.
* A set of hits for a single query may then be filtered with a caller-supplied filter, which will remove any
* alignments that do not pass the filter. If the primary alignment is removed, the best-mapping secondary alignment
* or alignment pair will be marked as primary.
*
*
* @throws IllegalStateException if the input is not queryname-sorted.
*/
class MultiHitAlignedReadIterator implements CloseableIterator<HitsForInsert> {
private final PeekableIterator<SAMRecord> peekIterator;
private final SAMRecordQueryNameComparator queryNameComparator = new SAMRecordQueryNameComparator();
private final PrimaryAlignmentSelectionStrategy primaryAlignmentSelectionStrategy;
private HitsForInsert theNext = null;
/**
*
* @param querynameOrderIterator
* @param primaryAlignmentSelectionStrategy Algorithm for selecting primary alignment when it is not clear from
* the input what should be primary.
*/
MultiHitAlignedReadIterator(final CloseableIterator<SAMRecord> querynameOrderIterator,
final PrimaryAlignmentSelectionStrategy primaryAlignmentSelectionStrategy) {
this.primaryAlignmentSelectionStrategy = primaryAlignmentSelectionStrategy;
peekIterator = new PeekableIterator<SAMRecord>(new FilteringIterator(querynameOrderIterator,
new SamRecordFilter() {
// Filter unmapped reads.
public boolean filterOut(final SAMRecord record) {
return record.getReadUnmappedFlag() || SAMUtils.cigarMapsNoBasesToRef(record.getCigar());
}
public boolean filterOut(final SAMRecord first, final SAMRecord second) {
return ((first.getReadUnmappedFlag() || SAMUtils.cigarMapsNoBasesToRef(first.getCigar()))
&& (second.getReadUnmappedFlag() || SAMUtils.cigarMapsNoBasesToRef(second.getCigar())));
}
}));
advance();
}
public void close() {
peekIterator.close();
}
public boolean hasNext() {
return theNext != null;
}
/**
* @throws IllegalStateException if the input is not queryname-sorted.
*/
public HitsForInsert next() {
if (!hasNext()) throw new NoSuchElementException();
final HitsForInsert ret = theNext;
advance();
return ret;
}
private void advance() {
while (peekIterator.hasNext()) {
theNext = nextMaybeEmpty();
if (theNext.numHits() > 0) return;
}
theNext = null;
}
private HitsForInsert nextMaybeEmpty() {
if (!peekIterator.hasNext()) throw new IllegalStateException();
final String readName = peekIterator.peek().getReadName();
final HitsForInsert hits = new HitsForInsert();
Boolean isPaired = null;
// Accumulate the alignments matching readName.
do {
final SAMRecord rec = peekIterator.next();
replaceHardWithSoftClips(rec);
// It is critical to do this here, because SamAlignmentMerger uses this exception to determine
// if the aligned input needs to be sorted.
if (peekIterator.hasNext() && queryNameComparator.fileOrderCompare(rec, peekIterator.peek()) > 0) {
throw new IllegalStateException("Underlying iterator is not queryname sorted: " +
rec + " > " + peekIterator.peek());
}
if (isPaired == null) {
isPaired = rec.getReadPairedFlag();
} else if (isPaired != rec.getReadPairedFlag()) {
throw new PicardException("Got a mix of paired and unpaired alignments for read " + readName);
}
// Records w/ a supplemental flag are stashed to the side until the primary alignment has
// been determined, and then re-added into the process later
if (!rec.getReadPairedFlag() || rec.getFirstOfPairFlag()) {
if (rec.getSupplementaryAlignmentFlag()) {
hits.addSupplementalFirstOfPairOrFragment(rec);
} else {
hits.addFirstOfPairOrFragment(rec);
}
} else if (rec.getSecondOfPairFlag()) {
if (rec.getSupplementaryAlignmentFlag()) {
hits.addSupplementalSecondOfPair(rec);
} else {
hits.addSecondOfPair(rec);
}
} else throw new PicardException("Read is marked as pair but neither first or second: " + readName);
} while (peekIterator.hasNext() && peekIterator.peek().getReadName().equals(readName));
// If there is no more than one alignment for each end, no need to do any coordination.
if (hits.numHits() <= 1) {
// No HI tags needed if only a single hit
if (hits.getFirstOfPair(0) != null) {
hits.getFirstOfPair(0).setAttribute(SAMTag.HI.name(), null);
hits.getFirstOfPair(0).setNotPrimaryAlignmentFlag(false);
}
if (hits.getSecondOfPair(0) != null) {
hits.getSecondOfPair(0).setAttribute(SAMTag.HI.name(), null);
hits.getSecondOfPair(0).setNotPrimaryAlignmentFlag(false);
}
} else {
primaryAlignmentSelectionStrategy.pickPrimaryAlignment(hits);
}
// Used to check that alignments for first and second were correlated, but this is no longer required.
return hits;
}
/** Replaces hard clips with soft clips and fills in bases and qualities with dummy values as needed. */
private void replaceHardWithSoftClips(final SAMRecord rec) {
if (rec.getReadUnmappedFlag()) return;
if (rec.getCigar().isEmpty()) return;
List<CigarElement> elements = rec.getCigar().getCigarElements();
final CigarElement first = elements.get(0);
final CigarElement last = elements.size() == 1 ? null : elements.get(elements.size()-1);
final int startHardClip = first.getOperator() == CigarOperator.H ? first.getLength() : 0;
final int endHardClip = (last != null && last.getOperator() == CigarOperator.H) ? last.getLength() : 0;
if (startHardClip + endHardClip > 0) {
final int len = rec.getReadBases().length + startHardClip + endHardClip;
// Fix the basecalls
final byte[] bases = new byte[len];
Arrays.fill(bases, (byte) 'N');
System.arraycopy(rec.getReadBases(), 0, bases, startHardClip, rec.getReadBases().length);
// Fix the quality scores
final byte[] quals = new byte[len];
Arrays.fill(quals, (byte) 2 );
System.arraycopy(rec.getBaseQualities(), 0, quals, startHardClip, rec.getBaseQualities().length);
// Fix the cigar!
elements = new ArrayList<CigarElement>(elements); // make it modifiable
if (startHardClip > 0) elements.set(0, new CigarElement(first.getLength(), CigarOperator.S));
if (endHardClip > 0) elements.set(elements.size()-1, new CigarElement(last.getLength(), CigarOperator.S));
// Set the update structures on the new record
rec.setReadBases(bases);
rec.setBaseQualities(quals);
rec.setCigar(new Cigar(elements));
}
}
/** Unsupported operation. */
public void remove() {
throw new UnsupportedOperationException();
}
}