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ref_aligner.h
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ref_aligner.h
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/*
* ref_aligner.h
*/
#ifndef REF_ALIGNER_H_
#define REF_ALIGNER_H_
#include <stdint.h>
#include <iostream>
#include <vector>
#include <stdexcept>
#include "seqan/sequence.h"
#include "alphabet.h"
#include "range.h"
#include "reference.h"
// Let the reference-aligner buffer size be 16K by default. If more
// room is required, a new buffer must be allocated from the heap.
const static int REF_ALIGNER_BUFSZ = 16 * 1024;
/**
* Abstract parent class for classes that look for alignments by
* matching against the reference sequence directly. This is useful
* both for sanity-checking results from the Bowtie index and for
* finding mates when the reference location of the opposite mate is
* known.
*/
template<typename TStr>
class RefAligner {
typedef seqan::String<seqan::Dna5> TDna5Str;
typedef seqan::String<char> TCharStr;
typedef std::vector<uint32_t> TU32Vec;
typedef std::vector<Range> TRangeVec;
typedef std::pair<uint64_t, uint64_t> TU64Pair;
typedef std::set<TU64Pair> TSetPairs;
public:
RefAligner(bool color,
bool verbose = false,
bool quiet = false,
uint32_t seedLen = 0,
uint32_t qualMax = 0xffffffff,
bool maqPenalty = false) :
color_(color), verbose_(verbose), seedLen_(seedLen),
qualMax_(qualMax), maqPenalty_(maqPenalty), refbuf_(buf_),
refbufSz_(REF_ALIGNER_BUFSZ), freeRefbuf_(false)
{ }
/**
* Free the reference-space alignment buffer if this object
* allocated it.
*/
virtual ~RefAligner() {
if(freeRefbuf_) {
delete[] refbuf_;
}
}
/**
* Find one alignment of qry:quals in the range begin-end in
* reference string ref. Store the alignment details in range.
*/
virtual void find(uint32_t numToFind,
const size_t tidx,
const BitPairReference *refs,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs = NULL,
TIndexOffU aoff = OFF_MASK,
bool seedOnLeft = false)
{
assert_gt(numToFind, 0);
assert_gt(end, begin);
TIndexOffU spread = end - begin + (color_ ? 1 : 0);
TIndexOffU spreadPlus = spread + 12;
// Make sure the buffer is large enough to accommodate the spread
if(spreadPlus > this->refbufSz_) {
this->newBuf(spreadPlus);
}
// Read in the relevant stretch of the reference string
int offset = refs->getStretch(this->refbuf_, tidx, begin, spread);
uint8_t *buf = ((uint8_t*)this->refbuf_) + offset;
if(color_) {
// Colorize buffer
for(size_t i = 0; i < (end-begin); i++) {
assert_leq((int)buf[i], 4);
buf[i] = dinuc2color[(int)buf[i]][(int)buf[i+1]];
}
}
// Look for alignments
ASSERT_ONLY(uint32_t irsz = (uint32_t)ranges.size());
anchor64Find(numToFind, tidx, buf, qry, quals, begin,
end, ranges, results, pairs, aoff, seedOnLeft);
#ifndef NDEBUG
for(size_t i = irsz; i < results.size(); i++) {
assert_eq(ranges[i].numMms, ranges[i].mms.size());
assert_eq(ranges[i].numMms, ranges[i].refcs.size());
}
#endif
}
/**
* Find one alignment of qry:quals in the range begin-end in
* reference string ref. Store the alignment details in range.
* Uses a combination of the anchor bases and 64-bit arithmetic to
* find anchors quickly.
*/
virtual void anchor64Find(uint32_t numToFind,
size_t tidx,
uint8_t* ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs = NULL,
TIndexOffU aoff = OFF_MASK,
bool seedOnLeft = false) const = 0;
/**
* Set a new reference-sequence buffer.
*/
void setBuf(uint32_t *newbuf, uint32_t newsz) {
if(freeRefbuf_) {
delete[] refbuf_;
freeRefbuf_ = false;
}
refbuf_ = newbuf;
refbufSz_ = newsz;
}
/**
* Set a new reference-sequence buffer.
*/
void newBuf(uint32_t newsz) {
if(freeRefbuf_) {
delete[] refbuf_;
}
try {
refbuf_ = new uint32_t[(newsz + 3) / 4];
if(refbuf_ == NULL) throw std::bad_alloc();
} catch(std::bad_alloc& e) {
cerr << "Error: Could not allocate reference-space alignment buffer of " << newsz << "B" << endl;
throw 1;
}
refbufSz_ = newsz;
freeRefbuf_ = true;
}
protected:
bool color_; /// whether to colorize reference buffers before handing off
bool verbose_; /// be talkative
uint32_t seedLen_; /// length of seed region for read
uint32_t qualMax_; /// maximum sum of quality penalties
bool maqPenalty_;/// whether to round as in Maq
uint32_t *refbuf_; /// pointer to current reference buffer
uint32_t refbufSz_; /// size of current reference buffer
uint32_t buf_[REF_ALIGNER_BUFSZ / 4]; /// built-in reference buffer (may be superseded)
bool freeRefbuf_; /// whether refbuf_ points to something we should delete
};
/**
* Concrete RefAligner for finding nearby exact hits given an anchor
* hit.
*/
template<typename TStr>
class ExactRefAligner : public RefAligner<TStr> {
typedef seqan::String<seqan::Dna5> TDna5Str;
typedef seqan::String<char> TCharStr;
typedef std::vector<uint32_t> TU32Vec;
typedef std::vector<Range> TRangeVec;
typedef std::pair<uint64_t, uint64_t> TU64Pair;
typedef std::set<TU64Pair> TSetPairs;
public:
ExactRefAligner(bool color, bool verbose, bool quiet) :
RefAligner<TStr>(color, verbose, quiet) { }
virtual ~ExactRefAligner() { }
protected:
/**
* Because we're doing end-to-end exact, we don't care which end of
* 'qry' is the 5' end.
*/
void naiveFind(uint32_t numToFind,
size_t tidx,
uint8_t* ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs,
TIndexOffU aoff,
bool seedOnLeft) const
{
assert_gt(numToFind, 0);
const uint32_t qlen = (uint32_t)seqan::length(qry);
assert_geq(end - begin, qlen); // caller should have checked this
assert_gt(end, begin);
assert_gt(qlen, 0);
TIndexOffU qend = end - qlen;
TIndexOffU lim = qend - begin;
TIndexOffU halfway = begin + (lim >> 1);
bool hi = false;
for(TIndexOffU i = 1; i <= lim+1; i++) {
TIndexOffU ri; // leftmost position in candidate alignment
TIndexOffU rir; // same, minus begin; for indexing into ref[]
if(hi) {
ri = halfway + (i >> 1); rir = ri - begin;
assert_leq(ri, qend);
} else {
ri = halfway - (i >> 1); rir = ri - begin;
assert_geq(ri, begin);
}
hi = !hi;
// Do the naive comparison
bool match = true;
for(size_t j = 0; j < qlen; j++) {
#if 0
// Count Ns in the reference as mismatches
const int q = (int)qry[j];
const int r = (int)ref[rir + j];
assert_leq(q, 4);
assert_leq(r, 4);
if(q == 4 || r == 4 || q != r) {
// Mismatch
match = false;
break;
}
// Match; continue
#else
// Disallow alignments that involve an N in the
// reference
const int r = (int)ref[rir + j];
if(r & 4) {
// N in reference; bail
match = false;
break;
}
const int q = (int)qry[j];
assert_leq(q, 4);
assert_lt(r, 4);
if(q != r) {
// Mismatch
match = false;
break;
}
// Match; continue
#endif
}
if(match) {
ranges.resize(ranges.size()+1);
Range& range = ranges.back();
range.stratum = 0;
range.numMms = 0;
assert_eq(0, range.mms.size());
assert_eq(0, range.refcs.size());
results.push_back(ri);
}
}
return; // no match
}
/**
* Because we're doing end-to-end exact, we don't care which end of
* 'qry' is the 5' end.
*/
virtual void anchor64Find(uint32_t numToFind,
size_t tidx,
uint8_t *ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs,
TIndexOffU aoff, // offset of anchor mate
bool seedOnLeft) const
{
assert_gt(numToFind, 0);
ASSERT_ONLY(const uint32_t rangesInitSz = (uint32_t)ranges.size());
ASSERT_ONLY(uint32_t duplicates = 0);
ASSERT_ONLY(uint32_t r2i = 0);
const uint32_t qlen = (uint32_t)seqan::length(qry);
assert_geq(end - begin, qlen); // caller should have checked this
assert_gt(end, begin);
assert_gt(qlen, 0);
#ifndef NDEBUG
// Get all naive hits
TRangeVec r2; std::vector<TIndexOffU> re2;
naiveFind(numToFind, tidx, ref, qry, quals, begin, end, r2,
re2, pairs, aoff, seedOnLeft);
#endif
const uint32_t anchorBitPairs = min<int>(qlen, 32);
// anchorOverhang = # read bases not included in the anchor
const TIndexOffU anchorOverhang = qlen <= 32 ? 0 : qlen - 32;
const TIndexOffU lim = end - qlen - begin;
const TIndexOffU halfway = begin + (lim >> 1);
uint64_t anchor = 0llu;
uint64_t buffw = 0llu;
// Set up a mask that we'll apply to the two bufs every round
// to discard bits that were rotated out of the anchor area
uint64_t clearMask = 0xffffffffffffffffllu;
bool useMask = false;
if(anchorBitPairs < 32) {
clearMask >>= ((32-anchorBitPairs) << 1);
useMask = true;
}
const int lhsShift = ((anchorBitPairs - 1) << 1);
// Build the contents of the 'anchor' dword and the initial
// contents of the 'buffw' dword. If there are fewer than 32
// anchorBitPairs, the content will be packed into the least
// significant bits of the word.
size_t skipLeftToRights = 0;
size_t skipRightToLefts = 0;
for(size_t i = 0; i < anchorBitPairs; i++) {
int c = (int)qry[i]; // next query character
assert_leq(c, 4);
if(c & 4) {
assert_eq(r2.size(), ranges.size() - rangesInitSz);
return; // can't match if query has Ns
}
int r = (int)ref[halfway - begin + i]; // next reference character
if(r & 4) {
r = 0;
// The right-to-left direction absorbs the candidate
// alignment based at 'halfway'; so skipLeftToRights is
// i, not i+1
skipLeftToRights = max(skipLeftToRights, i);
skipRightToLefts = max(skipRightToLefts, anchorBitPairs - i);
}
assert_lt(r, 4);
assert_lt(c, 4);
anchor = ((anchor << 2llu) | c);
buffw = ((buffw << 2llu) | r);
}
// Check whether read is disqualified by Ns outside of the anchor
// region
for(size_t i = anchorBitPairs; i < qlen; i++) {
if((int)qry[i] == 4) {
assert_eq(r2.size(), ranges.size() - rangesInitSz);
return; // can't match if query has Ns
}
}
uint64_t bufbw = buffw;
// We're moving the right-hand edge of the anchor along until
// it's 'anchorOverhang' chars from the end of the target region.
// Note that we're not making a 3'/5' distinction here; if we
// were, we might need to make the 'anchorOverhang' adjustment on
// the left end of the range rather than the right.
bool hi = false;
TIndexOffU riHi = halfway;
TIndexOffU rirHi = halfway - begin;
TIndexOffU rirHiAnchor = rirHi + anchorBitPairs - 1;
TIndexOffU riLo = halfway + 1;
TIndexOffU rirLo = halfway - begin + 1;
for(TIndexOffU i = 1; i <= lim + 1; i++) {
int r; // new reference char
assert_lt(skipLeftToRights, qlen);
assert_leq(skipRightToLefts, qlen);
if(hi) {
hi = false;
// Moving left-to-right
riHi++; rirHi++; rirHiAnchor++;
r = (int)ref[rirHiAnchor];
if(r & 4) {
r = 0;
skipLeftToRights = anchorBitPairs;
}
assert_lt(r, 4);
// Bring in new base pair at the least significant
// position
buffw = ((buffw << 2llu) | r);
if(useMask) buffw &= clearMask;
if(skipLeftToRights > 0) {
skipLeftToRights--;
continue;
}
if(buffw != anchor) {
continue;
}
} else {
hi = true;
// Moving right-to-left
riLo--; rirLo--;
r = (int)ref[rirLo];
if(r & 4) {
r = 0;
skipRightToLefts = qlen;
}
assert_lt(r, 4);
if(i >= 2) {
bufbw >>= 2llu;
// Bring in new base pair at the most significant
// position
bufbw |= ((uint64_t)r << lhsShift);
}
if(skipRightToLefts > 0) {
skipRightToLefts--;
continue;
}
if(bufbw != anchor) {
continue;
}
}
// Seed hit!
bool foundHit = true;
TIndexOffU ri = hi ? riLo : riHi;
TIndexOffU rir = hi ? rirLo : rirHi;
if(anchorOverhang > 0) {
// Does the non-anchor part of the alignment (the
// "overhang") ruin it?
bool skipCandidate = false;
for(size_t j = 0; j < anchorOverhang; j++) {
assert_lt(ri + anchorBitPairs + j, end);
int rc = (int)ref[rir + anchorBitPairs + j];
if(rc == 4) {
// Oops, encountered an N in the reference in
// the overhang portion of the candidate
// alignment
// (Note that we inverted hi earlier)
if(hi) {
// Right-to-left
skipRightToLefts = anchorOverhang - j - 1;
} else {
// Left-to-right
skipLeftToRights = anchorBitPairs + j;
}
skipCandidate = true;
break; // Skip this candidate
}
if((int)qry[32 + j] != rc) {
// Yes, overhang ruins it
foundHit = false;
break;
}
}
if(skipCandidate) continue;
}
if(foundHit) {
if(pairs != NULL) {
TU64Pair p;
if(ri < aoff) {
// By convention, the upstream mate's
// coordinates go in the 'first' field
p.first = ((uint64_t)tidx << 32) | (uint64_t)ri;
p.second = ((uint64_t)tidx << 32) | (uint64_t)aoff;
} else {
p.first = ((uint64_t)tidx << 32) | (uint64_t)aoff;
p.second = ((uint64_t)tidx << 32) | (uint64_t)ri;
}
if(pairs->find(p) != pairs->end()) {
// We already found this hit! Continue.
ASSERT_ONLY(duplicates++);
ASSERT_ONLY(r2i++);
continue;
} else {
// Record this hit
pairs->insert(p);
}
}
assert_lt(r2i, r2.size());
assert_eq(re2[r2i], ri);
ranges.resize(ranges.size()+1);
Range& range = ranges.back();
range.stratum = 0;
range.numMms = 0;
assert_eq(0, range.mms.size());
assert_eq(0, range.refcs.size());
ASSERT_ONLY(r2i++);
results.push_back(ri);
if(--numToFind == 0) return;
} else {
// Keep scanning
}
}
assert_leq(duplicates, r2.size());
assert_geq(r2.size() - duplicates, ranges.size() - rangesInitSz);
return; // no hit
}
};
/**
* Defined in ref_aligner.cpp. Maps an octet representing the XOR of
* two two-bit-per-base-encoded DNA sequences to the number of bases
* that mismatch between the two.
*/
extern unsigned char u8toMms[];
/**
* Concrete RefAligner for finding nearby 1-mismatch hits given an
* anchor hit.
*/
template<typename TStr>
class OneMMRefAligner : public RefAligner<TStr> {
typedef seqan::String<seqan::Dna5> TDna5Str;
typedef seqan::String<char> TCharStr;
typedef std::vector<Range> TRangeVec;
typedef std::vector<uint32_t> TU32Vec;
typedef std::pair<uint64_t, uint64_t> TU64Pair;
typedef std::set<TU64Pair> TSetPairs;
public:
OneMMRefAligner(bool color, bool verbose, bool quiet) :
RefAligner<TStr>(color, verbose, quiet) { }
virtual ~OneMMRefAligner() { }
protected:
/**
* Because we're doing end-to-end exact, we don't care which end of
* 'qry' is the 5' end.
*/
void naiveFind(uint32_t numToFind,
size_t tidx,
uint8_t* ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs,
TIndexOffU aoff,
bool seedOnLeft) const
{
assert_gt(numToFind, 0);
const uint32_t qlen = (uint32_t)seqan::length(qry);
assert_geq(end - begin, qlen); // caller should have checked this
assert_gt(end, begin);
assert_gt(qlen, 0);
TIndexOffU qend = end - qlen;
TIndexOffU lim = qend - begin;
TIndexOffU halfway = begin + (lim >> 1);
bool hi = false;
for(TIndexOffU i = 1; i <= lim+1; i++) {
TIndexOffU ri; // leftmost position in candidate alignment
TIndexOffU rir; // same, minus begin; for indexing into ref[]
if(hi) {
ri = halfway + (i >> 1); rir = ri - begin;
assert_leq(ri, qend);
} else {
ri = halfway - (i >> 1); rir = ri - begin;
assert_geq(ri, begin);
}
hi = !hi;
// Do the naive comparison
bool match = true;
int refc = -1;
TIndexOffU mmOff = OFF_MASK;
int mms = 0;
for(size_t j = 0; j < qlen; j++) {
#if 0
// Count Ns in the reference as mismatches
const int q = (int)qry[j];
const int r = (int)ref[rir + j];
assert_leq(q, 4);
assert_leq(r, 4);
if(q == 4 || r == 4 || q != r) {
#else
// Disallow alignments that involve an N in the
// reference
const int r = (int)ref[rir + j];
if(r & 4) {
// N in reference; bail
match = false;
break;
}
const int q = (int)qry[j];
assert_leq(q, 4);
assert_lt(r, 4);
if(q != r) {
#endif
// Mismatch!
if(++mms > 1) {
// Too many; reject this alignment
match = false;
break;
} else {
// First one; remember offset and ref char
refc = "ACGTN"[r];
mmOff = (uint32_t)j;
}
}
}
if(match) {
assert_leq(mms, 1);
ranges.resize(ranges.size()+1);
Range& range = ranges.back();
range.stratum = mms;
range.numMms = mms;
assert_eq(0, range.mms.size());
assert_eq(0, range.refcs.size());
if(mms == 1) {
assert_lt(mmOff, qlen);
range.mms.push_back(mmOff);
range.refcs.push_back(refc);
}
results.push_back(ri);
}
}
return;
}
/**
* Because we're doing end-to-end exact, we don't care which end of
* 'qry' is the 5' end.
*/
virtual void anchor64Find(uint32_t numToFind,
size_t tidx,
uint8_t* ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs = NULL,
TIndexOffU aoff = OFF_MASK,
bool seedOnLeft = false) const
{
assert_gt(numToFind, 0);
ASSERT_ONLY(const uint32_t rangesInitSz = (uint32_t)ranges.size());
ASSERT_ONLY(uint32_t duplicates = 0);
ASSERT_ONLY(uint32_t r2i = 0);
const uint32_t qlen = (uint32_t)seqan::length(qry);
assert_geq(end - begin, qlen); // caller should have checked this
assert_gt(end, begin);
assert_gt(qlen, 0);
#ifndef NDEBUG
// Get results from the naive matcher for sanity-checking
TRangeVec r2; std::vector<TIndexOffU> re2;
naiveFind(numToFind, tidx, ref, qry, quals, begin, end, r2,
re2, pairs, aoff, seedOnLeft);
#endif
const uint32_t anchorBitPairs = min<int>(qlen, 32);
const int lhsShift = ((anchorBitPairs - 1) << 1);
const uint32_t anchorCushion = 32 - anchorBitPairs;
// anchorOverhang = # read bases not included in the anchor
const size_t anchorOverhang = (qlen <= 32 ? 0 : (qlen - 32));
const TIndexOffU lim = end - qlen - begin;
const TIndexOffU halfway = begin + (lim >> 1);
uint64_t anchor = 0llu;
uint64_t buffw = 0llu; // rotating ref sequence buffer
// OR the 'diff' buffer with this so that we can always count
// 'N's as mismatches
uint64_t diffMask = 0llu;
// Set up a mask that we'll apply to the two bufs every round
// to discard bits that were rotated out of the anchor area
uint64_t clearMask = 0xffffffffffffffffllu;
bool useMask = false;
if(anchorBitPairs < 32) {
clearMask >>= ((32-anchorBitPairs) << 1);
useMask = true;
}
int nsInAnchor = 0;
int nPos = -1;
size_t skipLeftToRights = 0;
size_t skipRightToLefts = 0;
// Construct the 'anchor' 64-bit buffer so that it holds all of
// the first 'anchorBitPairs' bit pairs of the query.
for(size_t i = 0; i < anchorBitPairs; i++) {
int c = (int)qry[i]; // next query character
int r = (int)ref[halfway - begin + i]; // next reference character
if(r & 4) {
r = 0;
// The right-to-left direction absorbs the candidate
// alignment based at 'halfway'; so skipLeftToRights is
// i, not i+1
skipLeftToRights = max(skipLeftToRights, i);
skipRightToLefts = max(skipRightToLefts, anchorBitPairs - i);
}
assert_leq(c, 4);
assert_lt(r, 4);
// Special case: query has an 'N'
if(c == 4) {
if(++nsInAnchor > 1) {
// More than one 'N' in the anchor region; can't
// possibly have a 1-mismatch hit anywhere
assert_eq(r2.size(), ranges.size() - rangesInitSz);
return; // can't match if query has Ns
}
nPos = (int)i;
// Make it look like an 'A' in the anchor
c = 0;
diffMask = (diffMask << 2llu) | 1llu;
} else {
diffMask <<= 2llu;
}
anchor = ((anchor << 2llu) | c);
buffw = ((buffw << 2llu) | r);
}
// Check whether read is disqualified by Ns outside of the anchor
// region
for(size_t i = anchorBitPairs; i < qlen; i++) {
if((int)qry[i] == 4) {
if(++nsInAnchor > 1) {
assert_eq(r2.size(), ranges.size() - rangesInitSz);
return; // can't match if query has Ns
}
}
}
uint64_t bufbw = buffw;
// We're moving the right-hand edge of the anchor along until
// it's 'anchorOverhang' chars from the end of the target region.
// Note that we're not making a 3'/5' distinction here; if we
// were, we might need to make the 'anchorOverhang' adjustment on
// the left end of the range rather than the right.
bool hi = false;
TIndexOffU riHi = halfway;
TIndexOffU rirHi = halfway - begin;
TIndexOffU rirHiAnchor = rirHi + anchorBitPairs - 1;
TIndexOffU riLo = halfway + 1;
TIndexOffU rirLo = halfway - begin + 1;
for(TIndexOffU i = 1; i <= lim + 1; i++) {
int r; // new reference char
uint64_t diff;
assert_lt(skipLeftToRights, qlen);
assert_leq(skipRightToLefts, qlen);
if(hi) {
hi = false;
// Moving left-to-right
riHi++;
rirHi++;
rirHiAnchor++;
r = (int)ref[rirHiAnchor];
if(r & 4) {
r = 0;
skipLeftToRights = anchorBitPairs;
}
assert_lt(r, 4);
// Bring in new base pair at the least significant
// position
buffw = ((buffw << 2llu) | r);
if(useMask) buffw &= clearMask;
if(skipLeftToRights > 0) {
skipLeftToRights--;
continue;
}
diff = (buffw ^ anchor) | diffMask;
} else {
hi = true;
// Moving right-to-left
riLo--;
rirLo--;
r = (int)ref[rirLo];
if(r & 4) {
r = 0;
skipRightToLefts = qlen;
}
assert_lt(r, 4);
if(i >= 2) {
bufbw >>= 2llu;
// Bring in new base pair at the most significant
// position
bufbw |= ((uint64_t)r << lhsShift);
}
if(skipRightToLefts > 0) {
skipRightToLefts--;
continue;
}
diff = (bufbw ^ anchor) | diffMask;
}
if((diff & 0xffffffff00000000llu) &&
(diff & 0x00000000ffffffffllu)) continue;
TIndexOffU ri = hi ? riLo : riHi;
TIndexOffU rir = hi ? rirLo : rirHi;
// Could use pop count
uint8_t *diff8 = reinterpret_cast<uint8_t*>(&diff);
// As a first cut, see if there are too many mismatches in
// the first and last parts of the anchor
uint32_t diffs = u8toMms[(int)diff8[0]] + u8toMms[(int)diff8[7]];
if(diffs > 1) continue;
diffs += u8toMms[(int)diff8[1]] +
u8toMms[(int)diff8[2]] +
u8toMms[(int)diff8[3]] +
u8toMms[(int)diff8[4]] +
u8toMms[(int)diff8[5]] +
u8toMms[(int)diff8[6]];
uint32_t mmpos = 0xffffffff;
int refc = -1;
if(diffs > 1) {
// Too many differences
continue;
} else if(diffs == 1 && nPos != -1) {
// There was one difference, but there was also one N,
// so we already know where the difference is
mmpos = nPos;
refc = "ACGT"[(int)ref[rir + nPos]];
} else if(diffs == 1) {
// Figure out which position mismatched
mmpos = 31;
if((diff & 0xffffffffllu) == 0) { diff >>= 32llu; mmpos -= 16; }
assert_neq(0, diff);
if((diff & 0xffffllu) == 0) { diff >>= 16llu; mmpos -= 8; }
assert_neq(0, diff);
if((diff & 0xffllu) == 0) { diff >>= 8llu; mmpos -= 4; }
assert_neq(0, diff);
if((diff & 0xfllu) == 0) { diff >>= 4llu; mmpos -= 2; }
assert_neq(0, diff);
if((diff & 0x3llu) == 0) { mmpos--; }
assert_neq(0, diff);
assert_geq(mmpos, 0);
assert_lt(mmpos, 32);
mmpos -= anchorCushion;
refc = "ACGT"[(int)ref[rir + mmpos]];
}
// Now extend the anchor into a longer alignment
bool foundHit = true;
if(anchorOverhang > 0) {
assert_leq(ri + anchorBitPairs + anchorOverhang, end);
bool skipCandidate = false;
for(size_t j = 0; j < anchorOverhang; j++) {
int rc = (int)ref[rir + 32 + j];
if(rc == 4) {
// Oops, encountered an N in the reference in
// the overhang portion of the candidate
// alignment
// (Note that we inverted hi earlier)
if(hi) {
// Right-to-left
skipRightToLefts = anchorOverhang - j - 1;
} else {
// Left-to-right
skipLeftToRights = anchorBitPairs + j;
}
skipCandidate = true; // Skip this candidate
break;
}
if((int)qry[32 + j] != rc) {
if(++diffs > 1) {
foundHit = false;
break;
} else {
mmpos = (uint32_t)(32 + j);
refc = "ACGT"[(int)ref[rir + 32 + j]];
}
}
}
if(skipCandidate) continue;
}
if(!foundHit) continue;
if(pairs != NULL) {
TU64Pair p;
if(ri < aoff) {
// By convention, the upstream mate's
// coordinates go in the 'first' field
p.first = ((uint64_t)tidx << 32) | (uint64_t)ri;
p.second = ((uint64_t)tidx << 32) | (uint64_t)aoff;
} else {
p.second = ((uint64_t)tidx << 32) | (uint64_t)ri;
p.first = ((uint64_t)tidx << 32) | (uint64_t)aoff;
}
if(pairs->find(p) != pairs->end()) {
// We already found this hit! Continue.
ASSERT_ONLY(duplicates++);
ASSERT_ONLY(r2i++);
continue;
} else {
// Record this hit
pairs->insert(p);
}
}
assert_leq(diffs, 1);
assert_lt(r2i, r2.size());
assert_eq(re2[r2i], ri);
ranges.resize(ranges.size()+1);
Range& range = ranges.back();
assert_eq(diffs, r2[r2i].numMms);
range.stratum = diffs;
range.numMms = diffs;
assert_eq(0, range.mms.size());
assert_eq(0, range.refcs.size());
if(diffs == 1) {
assert_neq(mmpos, 0xffffffff);
assert_eq(mmpos, r2[r2i].mms[0]);
assert_neq(-1, refc);
assert_eq(refc, r2[r2i].refcs[0]);
range.mms.push_back(mmpos);
range.refcs.push_back(refc);
}
ASSERT_ONLY(r2i++);
results.push_back(ri);
if(--numToFind == 0) return;
}
assert_leq(duplicates, r2.size());
assert_geq(r2.size() - duplicates, ranges.size() - rangesInitSz);
return; // no hit
}
};
/**
* Concrete RefAligner for finding nearby 2-mismatch hits given an
* anchor hit.
*/
template<typename TStr>
class TwoMMRefAligner : public RefAligner<TStr> {
typedef seqan::String<seqan::Dna5> TDna5Str;
typedef seqan::String<char> TCharStr;
typedef std::vector<Range> TRangeVec;
typedef std::vector<uint32_t> TU32Vec;
typedef std::pair<uint64_t, uint64_t> TU64Pair;
typedef std::set<TU64Pair> TSetPairs;
public:
TwoMMRefAligner(bool color, bool verbose, bool quiet) :
RefAligner<TStr>(color, verbose, quiet) { }
virtual ~TwoMMRefAligner() { }
protected:
/**
* Because we're doing end-to-end exact, we don't care which end of
* 'qry' is the 5' end.
*/
void naiveFind(uint32_t numToFind,
size_t tidx,
uint8_t* ref,
const TDna5Str& qry,
const TCharStr& quals,
TIndexOffU begin,
TIndexOffU end,
TRangeVec& ranges,
std::vector<TIndexOffU>& results,
TSetPairs* pairs,
TIndexOffU aoff,
bool seedOnLeft) const
{
assert_gt(numToFind, 0);
const uint32_t qlen = (uint32_t)seqan::length(qry);
assert_geq(end - begin, qlen); // caller should have checked this
assert_gt(end, begin);
assert_gt(qlen, 0);
TIndexOffU qend = end - qlen;
TIndexOffU lim = qend - begin;
TIndexOffU halfway = begin + (lim >> 1);
bool hi = false;
for(TIndexOffU i = 1; i <= lim+1; i++) {
TIndexOffU ri; // leftmost position in candidate alignment
TIndexOffU rir; // same, minus begin; for indexing into ref[]
if(hi) {
ri = halfway + (i >> 1); rir = ri - begin;
assert_leq(ri, qend);
} else {
ri = halfway - (i >> 1); rir = ri - begin;
assert_geq(ri, begin);
}
hi = !hi;
// Do the naive comparison
bool match = true;
int refc1 = -1;
TIndexOffU mmOff1 = OFF_MASK;
int refc2 = -1;
TIndexOffU mmOff2 = OFF_MASK;
int mms = 0;
for(size_t j = 0; j < qlen; j++) {
#if 0
// Count Ns in the reference as mismatches
const int q = (int)qry[j];
const int r = (int)ref[rir + j];
assert_leq(q, 4);
assert_leq(r, 4);
if(q == 4 || r == 4 || q != r) {
#else
// Disallow alignments that involve an N in the
// reference
const int r = (int)ref[rir + j];
if(r & 4) {
// N in reference; bail
match = false;
break;
}
const int q = (int)qry[j];
assert_leq(q, 4);
assert_lt(r, 4);
if(q != r) {
#endif
// Mismatch!
if(++mms > 2) {
// Too many; reject this alignment
match = false;
break;
} else if(mms == 2) {
// Second one; remember offset and ref char