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/*
* The MIT License
*
* Copyright (c) 2009 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 htsjdk.samtools;
import htsjdk.samtools.util.PeekableIterator;
import java.util.Iterator;
import java.util.LinkedList;
import java.util.List;
import java.util.Queue;
/**
* Utility methods for pairs of SAMRecords
*/
public class SamPairUtil {
/**
* The possible orientations of paired reads.
*
* F = mapped to forward strand
* R = mapped to reverse strand
*
* FR means the read that's mapped to the forward strand comes before the
* read mapped to the reverse strand when their 5'-end coordinates are
* compared.
*/
public static enum PairOrientation
{
FR, // ( 5' --F--> <--R-- 5' ) - aka. innie
RF, // ( <--R-- 5' 5' --F--> ) - aka. outie
TANDEM // ( 5' --F--> 5' --F--> or ( <--R-- 5' <--R-- 5' )
}
/**
* Computes the pair orientation of the given SAMRecord.
* @param r
* @return PairOrientation of the given SAMRecord.
* @throws IllegalArgumentException If the record is not a paired read, or
* one or both reads are unmapped.
*/
public static PairOrientation getPairOrientation(final SAMRecord r)
{
final boolean readIsOnReverseStrand = r.getReadNegativeStrandFlag();
if(r.getReadUnmappedFlag() || !r.getReadPairedFlag() || r.getMateUnmappedFlag()) {
throw new IllegalArgumentException("Invalid SAMRecord: " + r.getReadName() + ". This method only works for SAMRecords " +
"that are paired reads with both reads aligned.");
}
if(readIsOnReverseStrand == r.getMateNegativeStrandFlag() ) {
return PairOrientation.TANDEM;
}
final long positiveStrandFivePrimePos = ( readIsOnReverseStrand
? r.getMateAlignmentStart() //mate's 5' position ( x---> )
: r.getAlignmentStart() ); //read's 5' position ( x---> )
final long negativeStrandFivePrimePos = ( readIsOnReverseStrand
? r.getAlignmentEnd() //read's 5' position ( <---x )
: r.getAlignmentStart() + r.getInferredInsertSize() ); //mate's 5' position ( <---x )
return ( positiveStrandFivePrimePos < negativeStrandFivePrimePos
? PairOrientation.FR
: PairOrientation.RF );
}
// TODO: KT and TF say this is more complicated than what I have here
public static boolean isProperPair(final SAMRecord firstEnd, final SAMRecord secondEnd,
final List expectedOrientations) {
// are both records mapped?
if (firstEnd.getReadUnmappedFlag() || secondEnd.getReadUnmappedFlag()) {
return false;
}
if (firstEnd.getReferenceName().equals(SAMRecord.NO_ALIGNMENT_REFERENCE_NAME)) {
return false;
}
// AND are they both mapped to the same chromosome
if (!firstEnd.getReferenceName().equals(secondEnd.getReferenceName())) {
return false;
}
// AND is the pair orientation in the set of expected orientations
final PairOrientation actual = getPairOrientation(firstEnd);
return expectedOrientations.contains(actual);
}
public static void assertMate(final SAMRecord firstOfPair, final SAMRecord secondOfPair) {
// Validate paired reads arrive as first of pair, then second of pair
if (firstOfPair == null) {
throw new SAMException(
"First record does not exist - cannot perform mate assertion!");
} else if (secondOfPair == null) {
throw new SAMException(
firstOfPair.toString() + " is missing its mate");
} else if (!firstOfPair.getReadPairedFlag()) {
throw new SAMException(
"First record is not marked as paired: " + firstOfPair.toString());
} else if (!secondOfPair.getReadPairedFlag()) {
throw new SAMException(
"Second record is not marked as paired: " + secondOfPair.toString());
} else if (!firstOfPair.getFirstOfPairFlag()) {
throw new SAMException(
"First record is not marked as first of pair: " + firstOfPair.toString());
} else if (!secondOfPair.getSecondOfPairFlag()) {
throw new SAMException(
"Second record is not marked as second of pair: " + secondOfPair.toString());
} else if (!firstOfPair.getReadName().equals(secondOfPair.getReadName())) {
throw new SAMException(
"First [" + firstOfPair.getReadName() + "] and Second [" +
secondOfPair.getReadName() + "] readnames do not match!");
}
}
/**
* Obtain the secondOfPair mate belonging to the firstOfPair SAMRecord
* (assumed to be in the next element of the specified samRecordIterator)
* @param samRecordIterator the iterator assumed to contain the secondOfPair SAMRecord in the
* next element in the iteration
* @param firstOfPair the firstOfPair SAMRecord
* @return the secondOfPair SAMRecord
* @throws SAMException when the secondOfPair mate cannot be obtained due to assertion failures
*/
public static SAMRecord obtainAssertedMate(final Iterator samRecordIterator,
final SAMRecord firstOfPair) {
if (samRecordIterator.hasNext()) {
final SAMRecord secondOfPair = samRecordIterator.next();
assertMate(firstOfPair, secondOfPair);
return secondOfPair;
} else {
throw new SAMException(
"Second record does not exist: " + firstOfPair.getReadName());
}
}
/**
* Compute SAMRecord insert size
* @param firstEnd
* @param secondEnd
* @return note that when storing insert size on the secondEnd, the return value must be negated.
*/
public static int computeInsertSize(final SAMRecord firstEnd, final SAMRecord secondEnd) {
if (firstEnd.getReadUnmappedFlag() || secondEnd.getReadUnmappedFlag()) {
return 0;
}
if (!firstEnd.getReferenceName().equals(secondEnd.getReferenceName())) {
return 0;
}
final int firstEnd5PrimePosition = firstEnd.getReadNegativeStrandFlag()? firstEnd.getAlignmentEnd(): firstEnd.getAlignmentStart();
final int secondEnd5PrimePosition = secondEnd.getReadNegativeStrandFlag()? secondEnd.getAlignmentEnd(): secondEnd.getAlignmentStart();
final int adjustment = (secondEnd5PrimePosition >= firstEnd5PrimePosition) ? +1 : -1;
return secondEnd5PrimePosition - firstEnd5PrimePosition + adjustment;
}
/**
* Write the mate info for two SAMRecords. This will always clear/remove any mate cigar tag that is present.
* @param rec1 the first SAM record
* @param rec2 the second SAM record
*/
public static void setMateInfo(final SAMRecord rec1, final SAMRecord rec2) {
setMateInfo(rec1, rec2, false);
}
/**
* Write the mate info for two SAMRecords
* @param rec1 the first SAM record. Must have a non-null SAMFileHeader.
* @param rec2 the second SAM record. Must have a non-null SAMFileHeader.
* @param setMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
*/
public static void setMateInfo(final SAMRecord rec1, final SAMRecord rec2, final boolean setMateCigar) {
// If neither read is unmapped just set their mate info
if (!rec1.getReadUnmappedFlag() && !rec2.getReadUnmappedFlag()) {
rec1.setMateReferenceIndex(rec2.getReferenceIndex());
rec1.setMateAlignmentStart(rec2.getAlignmentStart());
rec1.setMateNegativeStrandFlag(rec2.getReadNegativeStrandFlag());
rec1.setMateUnmappedFlag(false);
rec1.setAttribute(SAMTag.MQ, rec2.getMappingQuality());
rec2.setMateReferenceIndex(rec1.getReferenceIndex());
rec2.setMateAlignmentStart(rec1.getAlignmentStart());
rec2.setMateNegativeStrandFlag(rec1.getReadNegativeStrandFlag());
rec2.setMateUnmappedFlag(false);
rec2.setAttribute(SAMTag.MQ, rec1.getMappingQuality());
if (setMateCigar) {
rec1.setAttribute(SAMTag.MC, rec2.getCigarString());
rec2.setAttribute(SAMTag.MC, rec1.getCigarString());
}
else {
rec1.setAttribute(SAMTag.MC, null);
rec2.setAttribute(SAMTag.MC, null);
}
}
// Else if they're both unmapped set that straight
else if (rec1.getReadUnmappedFlag() && rec2.getReadUnmappedFlag()) {
rec1.setReferenceIndex(SAMRecord.NO_ALIGNMENT_REFERENCE_INDEX);
rec1.setAlignmentStart(SAMRecord.NO_ALIGNMENT_START);
rec1.setMateReferenceIndex(SAMRecord.NO_ALIGNMENT_REFERENCE_INDEX);
rec1.setMateAlignmentStart(SAMRecord.NO_ALIGNMENT_START);
rec1.setMateNegativeStrandFlag(rec2.getReadNegativeStrandFlag());
rec1.setMateUnmappedFlag(true);
rec1.setAttribute(SAMTag.MQ, null);
rec1.setAttribute(SAMTag.MC, null);
rec1.setInferredInsertSize(0);
rec2.setReferenceIndex(SAMRecord.NO_ALIGNMENT_REFERENCE_INDEX);
rec2.setAlignmentStart(SAMRecord.NO_ALIGNMENT_START);
rec2.setMateReferenceIndex(SAMRecord.NO_ALIGNMENT_REFERENCE_INDEX);
rec2.setMateAlignmentStart(SAMRecord.NO_ALIGNMENT_START);
rec2.setMateNegativeStrandFlag(rec1.getReadNegativeStrandFlag());
rec2.setMateUnmappedFlag(true);
rec2.setAttribute(SAMTag.MQ, null);
rec2.setAttribute(SAMTag.MC, null);
rec2.setInferredInsertSize(0);
}
// And if only one is mapped copy it's coordinate information to the mate
else {
final SAMRecord mapped = rec1.getReadUnmappedFlag() ? rec2 : rec1;
final SAMRecord unmapped = rec1.getReadUnmappedFlag() ? rec1 : rec2;
unmapped.setReferenceIndex(mapped.getReferenceIndex());
unmapped.setAlignmentStart(mapped.getAlignmentStart());
mapped.setMateReferenceIndex(unmapped.getReferenceIndex());
mapped.setMateAlignmentStart(unmapped.getAlignmentStart());
mapped.setMateNegativeStrandFlag(unmapped.getReadNegativeStrandFlag());
mapped.setMateUnmappedFlag(true);
mapped.setAttribute(SAMTag.MQ, null);
mapped.setAttribute(SAMTag.MC, null);
mapped.setInferredInsertSize(0);
unmapped.setMateReferenceIndex(mapped.getReferenceIndex());
unmapped.setMateAlignmentStart(mapped.getAlignmentStart());
unmapped.setMateNegativeStrandFlag(mapped.getReadNegativeStrandFlag());
unmapped.setMateUnmappedFlag(false);
unmapped.setAttribute(SAMTag.MQ, mapped.getMappingQuality());
// For the unmapped read, set mateCigar to the mate's Cigar, since the mate must be mapped
if (setMateCigar) unmapped.setAttribute(SAMTag.MC, mapped.getCigarString());
else unmapped.setAttribute(SAMTag.MC, null);
unmapped.setInferredInsertSize(0);
}
final int insertSize = SamPairUtil.computeInsertSize(rec1, rec2);
rec1.setInferredInsertSize(insertSize);
rec2.setInferredInsertSize(-insertSize);
}
/**
* Write the mate info for two SAMRecords
* @param rec1 the first SAM record
* @param rec2 the second SAM record
* @param header the SAM file header
* @param setMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
* @deprecated use {@link #setMateInfo(SAMRecord, SAMRecord, boolean)} instead
*/
@Deprecated
public static void setMateInfo(final SAMRecord rec1, final SAMRecord rec2, final SAMFileHeader header, final boolean setMateCigar) {
setMateInfo(rec1, rec2, setMateCigar);
}
/**
* Write the mate info for two SAMRecords. This will always clear/remove any mate cigar tag that is present.
* @param rec1 the first SAM record
* @param rec2 the second SAM record
* @param header the SAM file header
* @deprecated use {@link #setMateInfo(SAMRecord, SAMRecord)} instead
*/
@Deprecated
public static void setMateInfo(final SAMRecord rec1, final SAMRecord rec2, final SAMFileHeader header) {
setMateInfo(rec1, rec2);
}
/**
* Sets mate pair information appropriately on a supplemental SAMRecord (e.g. from a split alignment)
* using the primary alignment of the read's mate.
* @param supplemental a supplemental alignment for the mate pair of the primary supplied
* @param matePrimary the primary alignment of the the mate pair of the supplemental
* @param setMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
*/
public static void setMateInformationOnSupplementalAlignment( final SAMRecord supplemental,
final SAMRecord matePrimary,
final boolean setMateCigar) {
supplemental.setMateReferenceIndex(matePrimary.getReferenceIndex());
supplemental.setMateAlignmentStart(matePrimary.getAlignmentStart());
supplemental.setMateNegativeStrandFlag(matePrimary.getReadNegativeStrandFlag());
supplemental.setMateUnmappedFlag(matePrimary.getReadUnmappedFlag());
supplemental.setInferredInsertSize(-matePrimary.getInferredInsertSize());
if (setMateCigar && !matePrimary.getReadUnmappedFlag()) {
supplemental.setAttribute(SAMTag.MC, matePrimary.getCigarString());
}
else {
supplemental.setAttribute(SAMTag.MC, null);
}
}
/**
* Sets mate pair information appropriately on a supplemental SAMRecord (e.g. from a split alignment)
* using the primary alignment of the read's mate.
* @param supplemental a supplemental alignment for the mate pair of the primary supplied
* @param matePrimary the primary alignment of the the mate pair of the supplemental
*/
public static void setMateInformationOnSupplementalAlignment( final SAMRecord supplemental,
final SAMRecord matePrimary) {
setMateInformationOnSupplementalAlignment(supplemental, matePrimary, false);
}
/**
* This method will clear any mate cigar already present.
* @deprecated use {@link #setProperPairAndMateInfo(SAMRecord, SAMRecord, List)} instead
*/
@Deprecated
public static void setProperPairAndMateInfo(final SAMRecord rec1, final SAMRecord rec2,
final SAMFileHeader header,
final List expectedOrientations) {
setProperPairAndMateInfo(rec1, rec2, expectedOrientations);
}
/**
* @param addMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
* @deprecated use {@link #setProperPairAndMateInfo(SAMRecord, SAMRecord, List, boolean)}
*/
@Deprecated
public static void setProperPairAndMateInfo(final SAMRecord rec1, final SAMRecord rec2,
final SAMFileHeader header,
final List expectedOrientations,
final boolean addMateCigar) {
setProperPairAndMateInfo(rec1, rec2, expectedOrientations, addMateCigar);
}
/**
* This method will clear any mate cigar already present.
*/
public static void setProperPairAndMateInfo(final SAMRecord rec1, final SAMRecord rec2,
final List expectedOrientations) {
setProperPairAndMateInfo(rec1, rec2, expectedOrientations, false);
}
/**
* @param addMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
*/
public static void setProperPairAndMateInfo(final SAMRecord rec1, final SAMRecord rec2,
final List expectedOrientations,
final boolean addMateCigar) {
setMateInfo(rec1, rec2, addMateCigar);
setProperPairFlags(rec1, rec2, expectedOrientations);
}
public static void setProperPairFlags(final SAMRecord rec1, final SAMRecord rec2, final List expectedOrientations) {
final boolean properPair = (!rec1.getReadUnmappedFlag() && !rec2.getReadUnmappedFlag())
? isProperPair(rec1, rec2, expectedOrientations)
: false;
rec1.setProperPairFlag(properPair);
rec2.setProperPairFlag(properPair);
}
/**
* A class to iterate through SAMRecords and set mate information on the given records, and optionally
* set the mate cigar tag (true by default).
*/
public static class SetMateInfoIterator extends PeekableIterator {
private final Queue records = new LinkedList();
private final boolean setMateCigar;
private final boolean ignoreMissingMates;
private long numMateCigarsAdded = 0;
/**
* By default, the mate cigar tag is set
* @param iterator the iterator to wrap
*/
public SetMateInfoIterator(final Iterator iterator) {
this(iterator, true);
}
/**
* @param iterator the iterator to wrap
* @param setMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
*/
public SetMateInfoIterator(final Iterator iterator, final boolean setMateCigar) {
this(iterator, setMateCigar, false);
}
/**
* @param iterator the iterator to wrap
* @param setMateCigar true if we are to update/create the Mate CIGAR (MC) optional tag, false if we are to clear any mate cigar tag that is present.
* @param ignoreMissingMates set this to true if we are to ignore missing mates, otherwise an exception will be thrown when a missing mate is encountered
*/
public SetMateInfoIterator(final Iterator iterator, final boolean setMateCigar, final boolean ignoreMissingMates) {
super(iterator);
this.setMateCigar = setMateCigar;
this.ignoreMissingMates = ignoreMissingMates;
}
/**
* @return the current number of mate cigars added. This could be more than the number of records returned.
*/
public long getNumMateCigarsAdded() { return this.numMateCigarsAdded; }
@Override
public boolean hasNext() {
return (!records.isEmpty() || super.hasNext());
}
/**
* Populate this.records if necessary.
*/
private void advance() {
// No need to advance if we have records remaining
if (!records.isEmpty()) return;
/*
Get all records with the same name, and then identify the canonical first and second end to which we
want to set mate info.
*/
SAMRecord firstPrimaryRecord = null, secondPrimaryRecord = null;
final SAMRecord first = super.peek(); // peek so we consider it in the following loop
boolean containsSupplementalRecord = false;
while (super.hasNext() && super.peek().getReadName().equals(first.getReadName())) {
final SAMRecord record = super.next();
// We must make sure that we find only one "primary" alignments for each end
if (record.getReadPairedFlag()) {
if (!record.isSecondaryOrSupplementary()) {
if (record.getFirstOfPairFlag()) {
if (null != firstPrimaryRecord) {
throw new SAMException("Found two records that are paired, not supplementary, and first of the pair: " + record.getReadName());
}
firstPrimaryRecord = record;
} else if (record.getSecondOfPairFlag()) {
if (null != secondPrimaryRecord) {
throw new SAMException("Found two records that are paired, not supplementary, and second of the pair: " + record.getReadName());
}
secondPrimaryRecord = record;
}
}
if (record.getSupplementaryAlignmentFlag()) containsSupplementalRecord = true;
}
records.add(record);
}
// TODO: should we check that we do not have a mix of paired and fragment reads?
// we must find both records to update the mate info
if (null != firstPrimaryRecord && null != secondPrimaryRecord) {
// Update mate info
SamPairUtil.setMateInfo(firstPrimaryRecord, secondPrimaryRecord, this.setMateCigar);
if (this.setMateCigar) this.numMateCigarsAdded += 2;
// Set mate information on supplemental records
if (containsSupplementalRecord) {
for (final SAMRecord record : records) {
if (record.getReadPairedFlag() && record.getSupplementaryAlignmentFlag()) {
if (record.getFirstOfPairFlag()) {
SamPairUtil.setMateInformationOnSupplementalAlignment(record, secondPrimaryRecord, this.setMateCigar);
} else {
SamPairUtil.setMateInformationOnSupplementalAlignment(record, firstPrimaryRecord, this.setMateCigar);
}
this.numMateCigarsAdded++;
}
}
}
} else if (!this.ignoreMissingMates) {
if (null != firstPrimaryRecord && firstPrimaryRecord.getReadPairedFlag()) {
throw new SAMException("Missing second read of pair: " + firstPrimaryRecord.getReadName());
} else if (null != secondPrimaryRecord && secondPrimaryRecord.getReadPairedFlag()) {
throw new SAMException("Missing first read of pair: " + secondPrimaryRecord.getReadName());
}
}
}
@Override
public SAMRecord next() {
advance();
if (records.isEmpty()) throw new IllegalStateException("Unexpectedly found an empty record list");
return this.records.poll();
}
@Override
public SAMRecord peek() {
advance();
if (records.isEmpty()) throw new IllegalStateException("Unexpectedly found an empty record list");
return this.records.peek();
}
}
}
