private GATKSAMRecord revertSoftClippedBases(GATKSAMRecord read) {
GATKSAMRecord unclipped = (GATKSAMRecord) read.clone();
Cigar unclippedCigar = new Cigar();
int matchesCount = 0;
for (CigarElement element : read.getCigar().getCigarElements()) {
if (element.getOperator() == CigarOperator.SOFT_CLIP
|| element.getOperator() == CigarOperator.MATCH_OR_MISMATCH)
matchesCount += element.getLength();
else if (matchesCount > 0) {
unclippedCigar.add(new CigarElement(matchesCount, CigarOperator.MATCH_OR_MISMATCH));
matchesCount = 0;
unclippedCigar.add(element);
} else unclippedCigar.add(element);
}
if (matchesCount > 0)
unclippedCigar.add(new CigarElement(matchesCount, CigarOperator.MATCH_OR_MISMATCH));
unclipped.setCigar(unclippedCigar);
final int newStart =
read.getAlignmentStart() + calculateAlignmentStartShift(read.getCigar(), unclippedCigar);
unclipped.setAlignmentStart(newStart);
if (newStart <= 0) {
// if the start of the unclipped read occurs before the contig,
// we must hard clip away the bases since we cannot represent reads with
// negative or 0 alignment start values in the SAMRecord (e.g., 0 means unaligned)
return hardClip(unclipped, 0, -newStart);
} else {
return unclipped;
}
}
@Override
protected Double getElementForRead(final GATKSAMRecord read, final int refLoc) {
final int offset =
ReadUtils.getReadCoordinateForReferenceCoordinate(
read.getSoftStart(), read.getCigar(), refLoc, ReadUtils.ClippingTail.RIGHT_TAIL, true);
if (offset == ReadUtils.CLIPPING_GOAL_NOT_REACHED) return null;
int readPos = AlignmentUtils.calcAlignmentByteArrayOffset(read.getCigar(), offset, false, 0, 0);
final int numAlignedBases = AlignmentUtils.getNumAlignedBasesCountingSoftClips(read);
if (readPos > numAlignedBases / 2) readPos = numAlignedBases - (readPos + 1);
return (double) readPos;
}
/**
* Hard clip bases from read, from start to stop in base coordinates
*
* <p>If start == 0, then we will clip from the front of the read, otherwise we clip from the
* right. If start == 0 and stop == 10, this would clip out the first 10 bases of the read.
*
* <p>Note that this function works with reads with negative alignment starts, in order to allow
* us to hardClip reads that have had their soft clips reverted and so might have negative
* alignment starts
*
* <p>Works properly with reduced reads and insertion/deletion base qualities
*
* @param read a non-null read
* @param start a start >= 0 and < read.length
* @param stop a stop >= 0 and < read.length.
* @return a cloned version of read that has been properly trimmed down
*/
private GATKSAMRecord hardClip(GATKSAMRecord read, int start, int stop) {
// If the read is unmapped there is no Cigar string and neither should we create a new cigar
// string
final CigarShift cigarShift =
(read.getReadUnmappedFlag())
? new CigarShift(new Cigar(), 0, 0)
: hardClipCigar(read.getCigar(), start, stop);
// the cigar may force a shift left or right (or both) in case we are left with insertions
// starting or ending the read after applying the hard clip on start/stop.
final int newLength =
read.getReadLength()
- (stop - start + 1)
- cigarShift.shiftFromStart
- cigarShift.shiftFromEnd;
final byte[] newBases = new byte[newLength];
final byte[] newQuals = new byte[newLength];
final int copyStart =
(start == 0) ? stop + 1 + cigarShift.shiftFromStart : cigarShift.shiftFromStart;
System.arraycopy(read.getReadBases(), copyStart, newBases, 0, newLength);
System.arraycopy(read.getBaseQualities(), copyStart, newQuals, 0, newLength);
final GATKSAMRecord hardClippedRead = (GATKSAMRecord) read.clone();
hardClippedRead
.resetSoftStartAndEnd(); // reset the cached soft start and end because they may have
// changed now that the read was hard clipped. No need to calculate
// them now. They'll be lazily calculated on the next call to
// getSoftStart()/End()
hardClippedRead.setBaseQualities(newQuals);
hardClippedRead.setReadBases(newBases);
hardClippedRead.setCigar(cigarShift.cigar);
if (start == 0)
hardClippedRead.setAlignmentStart(
read.getAlignmentStart()
+ calculateAlignmentStartShift(read.getCigar(), cigarShift.cigar));
if (read.hasBaseIndelQualities()) {
final byte[] newBaseInsertionQuals = new byte[newLength];
final byte[] newBaseDeletionQuals = new byte[newLength];
System.arraycopy(
read.getBaseInsertionQualities(), copyStart, newBaseInsertionQuals, 0, newLength);
System.arraycopy(
read.getBaseDeletionQualities(), copyStart, newBaseDeletionQuals, 0, newLength);
hardClippedRead.setBaseQualities(newBaseInsertionQuals, EventType.BASE_INSERTION);
hardClippedRead.setBaseQualities(newBaseDeletionQuals, EventType.BASE_DELETION);
}
return hardClippedRead;
}
private List<PileupElement> makeReads(final int n, final int mapq, final String op) {
final int readLength = 3;
final List<PileupElement> elts = new LinkedList<PileupElement>();
for (int i = 0; i < n; i++) {
GATKSAMRecord read =
ArtificialSAMUtils.createArtificialRead(header, "read", 0, 1, readLength);
read.setReadBases(Utils.dupBytes((byte) 'A', readLength));
read.setBaseQualities(Utils.dupBytes((byte) 30, readLength));
read.setCigarString("1M1" + op + "1M");
read.setMappingQuality(mapq);
final int baseOffset = op.equals("M") ? 1 : 0;
final CigarElement cigarElement = read.getCigar().getCigarElement(1);
elts.add(new PileupElement(read, baseOffset, cigarElement, 1, 0));
}
return elts;
}
@Test
public void testRBPMappingQuals() {
// create a read with high MQ
final GATKSAMRecord read = ArtificialSAMUtils.createArtificialRead(header, "read", 0, 1, 10);
read.setReadBases(Utils.dupBytes((byte) 'A', 10));
read.setBaseQualities(Utils.dupBytes((byte) 30, 10));
read.setCigarString("10M");
read.setMappingQuality(200); // set a MQ higher than max signed byte
// now create the RBP
final List<PileupElement> elts = new LinkedList<>();
elts.add(new PileupElement(read, 0, read.getCigar().getCigarElement(0), 0, 0));
final Map<String, ReadBackedPileupImpl> pileupsBySample = new HashMap<>();
pileupsBySample.put("foo", new ReadBackedPileupImpl(loc, elts));
final ReadBackedPileup pileup = new ReadBackedPileupImpl(loc, pileupsBySample);
Assert.assertEquals(pileup.getMappingQuals()[0], 200);
}
/**
* @param read a read containing the variant
* @return number of hard clipped and low qual bases at the read end (where end is right end
* w.r.t. the reference)
*/
public static int getNumClippedBasesAtEnd(final GATKSAMRecord read) {
// check for hard clips (never consider these bases):
final Cigar c = read.getCigar();
CigarElement last = c.getCigarElement(c.numCigarElements() - 1);
int numEndClippedBases = 0;
if (last.getOperator() == CigarOperator.H) {
numEndClippedBases = last.getLength();
}
final byte[] unclippedReadBases = read.getReadBases();
final byte[] unclippedReadQuals = read.getBaseQualities();
// Do a stricter base clipping than provided by CIGAR string, since this one may be too
// conservative,
// and may leave a string of Q2 bases still hanging off the reads.
// TODO: this code may not even get used because HaplotypeCaller already hard clips low quality
// tails
for (int i = unclippedReadBases.length - numEndClippedBases - 1; i >= 0; i--) {
if (unclippedReadQuals[i] < PairHMMIndelErrorModel.BASE_QUAL_THRESHOLD) numEndClippedBases++;
else break;
}
return numEndClippedBases;
}
/**
* Clips the bases in read according to this operation's start and stop. Uses the clipping
* representation used is the one provided by algorithm argument.
*
* @param algorithm clipping algorithm to use
* @param originalRead the read to be clipped
*/
public GATKSAMRecord apply(ClippingRepresentation algorithm, GATKSAMRecord originalRead) {
GATKSAMRecord read = (GATKSAMRecord) originalRead.clone();
byte[] quals = read.getBaseQualities();
byte[] bases = read.getReadBases();
byte[] newBases = new byte[bases.length];
byte[] newQuals = new byte[quals.length];
switch (algorithm) {
// important note:
// it's not safe to call read.getReadBases()[i] = 'N' or read.getBaseQualities()[i] = 0
// because you're not guaranteed to get a pointer to the actual array of bytes in the
// GATKSAMRecord
case WRITE_NS:
for (int i = 0; i < bases.length; i++) {
if (i >= start && i <= stop) {
newBases[i] = 'N';
} else {
newBases[i] = bases[i];
}
}
read.setReadBases(newBases);
break;
case WRITE_Q0S:
for (int i = 0; i < quals.length; i++) {
if (i >= start && i <= stop) {
newQuals[i] = 0;
} else {
newQuals[i] = quals[i];
}
}
read.setBaseQualities(newQuals);
break;
case WRITE_NS_Q0S:
for (int i = 0; i < bases.length; i++) {
if (i >= start && i <= stop) {
newQuals[i] = 0;
newBases[i] = 'N';
} else {
newQuals[i] = quals[i];
newBases[i] = bases[i];
}
}
read.setBaseQualities(newBases);
read.setReadBases(newBases);
break;
case HARDCLIP_BASES:
read = hardClip(read, start, stop);
break;
case SOFTCLIP_BASES:
if (read.getReadUnmappedFlag()) {
// we can't process unmapped reads
throw new UserException("Read Clipper cannot soft clip unmapped reads");
}
// System.out.printf("%d %d %d%n", stop, start, read.getReadLength());
int myStop = stop;
if ((stop + 1 - start) == read.getReadLength()) {
// BAM representation issue -- we can't SOFTCLIP away all bases in a read, just leave it
// alone
// Walker.logger.info(String.format("Warning, read %s has all bases clip but this can't be
// represented with SOFTCLIP_BASES, just leaving it alone", read.getReadName()));
// break;
myStop--; // just decrement stop
}
if (start > 0 && myStop != read.getReadLength() - 1)
throw new RuntimeException(
String.format(
"Cannot apply soft clipping operator to the middle of a read: %s to be clipped at %d-%d",
read.getReadName(), start, myStop));
Cigar oldCigar = read.getCigar();
int scLeft = 0, scRight = read.getReadLength();
if (start == 0) scLeft = myStop + 1;
else scRight = start;
Cigar newCigar = softClip(oldCigar, scLeft, scRight);
read.setCigar(newCigar);
int newClippedStart = getNewAlignmentStartOffset(newCigar, oldCigar);
int newStart = read.getAlignmentStart() + newClippedStart;
read.setAlignmentStart(newStart);
break;
case REVERT_SOFTCLIPPED_BASES:
read = revertSoftClippedBases(read);
break;
default:
throw new IllegalStateException("Unexpected Clipping operator type " + algorithm);
}
return read;
}
@Override
protected boolean isUsableRead(final GATKSAMRecord read, final int refLoc) {
return super.isUsableRead(read, refLoc)
&& read.getSoftStart() + read.getCigar().getReadLength() > refLoc;
}