static int aux_fields1(void)
{
static const char sam[] = "data:"
"@SQ\tSN:one\tLN:1000\n"
"@SQ\tSN:two\tLN:500\n"
"r1\t0\tone\t500\t20\t8M\t*\t0\t0\tATGCATGC\tqqqqqqqq\tXA:A:k\tXi:i:37\tXf:f:" xstr(PI) "\tXd:d:" xstr(E) "\tXZ:Z:" HELLO "\tXH:H:" BEEF "\tXB:B:c,-2,0,+2\tZZ:i:1000000\n";
// Canonical form of the alignment record above, as output by sam_format1()
static const char r1[] = "r1\t0\tone\t500\t20\t8M\t*\t0\t0\tATGCATGC\tqqqqqqqq\tXA:A:k\tXi:i:37\tXf:f:3.14159\tXd:d:2.71828\tXZ:Z:" HELLO "\tXH:H:" BEEF "\tXB:B:c,-2,0,2\tZZ:i:1000000";
samFile *in = sam_open(sam, "r");
bam_hdr_t *header = sam_hdr_read(in);
bam1_t *aln = bam_init1();
uint8_t *p;
uint32_t n;
kstring_t ks = { 0, 0, NULL };
if (sam_read1(in, header, aln) >= 0) {
if ((p = check_bam_aux_get(aln, "XA", 'A')) && bam_aux2A(p) != 'k')
fail("XA field is '%c', expected 'k'", bam_aux2A(p));
if ((p = check_bam_aux_get(aln, "Xi", 'C')) && bam_aux2i(p) != 37)
fail("Xi field is %d, expected 37", bam_aux2i(p));
if ((p = check_bam_aux_get(aln, "Xf", 'f')) && fabs(bam_aux2f(p) - PI) > 1E-6)
fail("Xf field is %.12f, expected pi", bam_aux2f(p));
if ((p = check_bam_aux_get(aln, "Xd", 'd')) && fabs(bam_aux2f(p) - E) > 1E-6)
fail("Xf field is %.12f, expected e", bam_aux2f(p));
if ((p = check_bam_aux_get(aln, "XZ", 'Z')) && strcmp(bam_aux2Z(p), HELLO) != 0)
fail("XZ field is \"%s\", expected \"%s\"", bam_aux2Z(p), HELLO);
if ((p = check_bam_aux_get(aln, "XH", 'H')) && strcmp(bam_aux2Z(p), BEEF) != 0)
fail("XH field is \"%s\", expected \"%s\"", bam_aux2Z(p), BEEF);
// TODO Invent and use bam_aux2B()
if ((p = check_bam_aux_get(aln, "XB", 'B')) && ! (memcmp(p, "Bc", 2) == 0 && (memcpy(&n, p+2, 4), n) == 3 && memcmp(p+6, "\xfe\x00\x02", 3) == 0))
fail("XB field is %c,..., expected c,-2,0,+2", p[1]);
if ((p = check_bam_aux_get(aln, "ZZ", 'I')) && bam_aux2i(p) != 1000000)
fail("ZZ field is %d, expected 1000000", bam_aux2i(p));
if (sam_format1(header, aln, &ks) < 0)
fail("can't format record");
if (strcmp(ks.s, r1) != 0)
fail("record formatted incorrectly: \"%s\"", ks.s);
free(ks.s);
}
else fail("can't read record");
bam_destroy1(aln);
bam_hdr_destroy(header);
sam_close(in);
return 1;
}
static int test_update_array(bam1_t *aln, const char target_id[2],
uint8_t type, uint32_t nitems, void *data,
const char next_id[2], int64_t next_val,
char next_type)
{
uint8_t *p;
// Try updating target
if (bam_aux_update_array(aln, target_id, type, nitems, data) < 0) {
fail("update %2.s tag", target_id);
return -1;
}
// Check values
p = bam_aux_get(aln, target_id);
if (!p) {
fail("find %.2s tag", target_id);
return -1;
}
switch (type) {
case 'c':
CHECK_ARRAY_VALS(int8_t, bam_auxB2i, PRId64, PRId8); break;
case 'C':
CHECK_ARRAY_VALS(uint8_t, bam_auxB2i, PRId64, PRIu8); break;
case 's':
CHECK_ARRAY_VALS(int16_t, bam_auxB2i, PRId64, PRId16); break;
case 'S':
CHECK_ARRAY_VALS(uint16_t, bam_auxB2i, PRId64, PRIu16); break;
case 'i':
CHECK_ARRAY_VALS(int32_t, bam_auxB2i, PRId64, PRId32); break;
case 'I':
CHECK_ARRAY_VALS(uint32_t, bam_auxB2i, PRId64, PRIu32); break;
case 'f':
CHECK_ARRAY_VALS(float, bam_auxB2f, "e", "e"); break;
}
// If given, check that the next tag hasn't been clobbered by the
// update above.
if (!*next_id) return 0;
p = bam_aux_get(aln, next_id);
if (!p) {
fail("find %.2s tag after updating %.2s", next_id, target_id);
return -1;
}
if (*p != next_type || bam_aux2i(p) != next_val) {
fail("after updating %.2s:"
" %.2s field is %c:%"PRId64"; expected %c:%"PRId64,
target_id, next_id, *p, bam_aux2i(p), next_type, next_val);
return -1;
}
return 0;
}
int32_t getMismatches() const
{
assert(m_dataPtr);
uint8_t *mm = bam_aux_get(m_dataPtr.get(), "NM");
if (mm != NULL)
return bam_aux2i(mm);
return NO_COLOR_MM;
}
int getIndelAmbiguity() const
{
assert(m_dataPtr);
uint8_t *amb = bam_aux_get(m_dataPtr.get(), "XA");
if (amb != NULL)
return bam_aux2i(amb);
return INDEL_NO_AMBIGUITY;
}
bool isMappedUnique() const
{
assert(m_dataPtr);
uint8_t *hits = bam_aux_get(m_dataPtr.get(), "NH");
if (hits != NULL)
return (bam_aux2i(hits) == 1);
return false;
}
int32_t getReportedAlignments() const
{
assert(m_dataPtr);
uint8_t *hits = bam_aux_get(m_dataPtr.get(), "NH");
if (hits != NULL)
return bam_aux2i(hits);
return NO_NH;
}
int32_t getMapQual() const // comes from SM tag, applies to this read if this read has a mate.
{
assert(m_dataPtr);
//assert(shouldHaveMate()); // This will be controversial!
uint8_t *mq = bam_aux_get(m_dataPtr.get(), "SM");
if (mq != NULL)
return bam_aux2i(mq);
return NO_MAP_QUAL;
}
int32_t
tmap_sam_get_fo_start_idx(tmap_sam_t *sam)
{
uint8_t *tag = NULL;
// ZF
tag = bam_aux_get(sam->b, "ZF");
if(NULL != tag) return bam_aux2i(tag);
else return -1;
}
int32_t
tmap_sam_get_za(tmap_sam_t *sam)
{
uint8_t *tag = NULL;
// ZA
if(NULL == sam->b) tmap_bug();
tag = bam_aux_get(sam->b, "ZA");
if(NULL != tag) return bam_aux2i(tag);
else return -1;
}
static int test_update_int(bam1_t *aln,
const char target_id[2], int64_t target_val,
char expected_type,
const char next_id[2], int64_t next_val,
char next_type) {
uint8_t *p;
// Try updating target
if (bam_aux_update_int(aln, target_id, target_val) < 0) {
fail("update %.2s tag", target_id);
return -1;
}
// Check it's there and has the right type and value
p = bam_aux_get(aln, target_id);
if (!p) {
fail("find %.2s tag", target_id);
return -1;
}
if (*p != expected_type || bam_aux2i(p) != target_val) {
fail("%.2s field is %c:%"PRId64"; expected %c:%"PRId64,
target_id, *p, bam_aux2i(p), expected_type, target_val);
return -1;
}
// If given, check that the next tag hasn't been clobbered by the
// update above.
if (!*next_id) return 0;
p = bam_aux_get(aln, next_id);
if (!p) {
fail("find %.2s tag after updating %.2s", next_id, target_id);
return -1;
}
if (*p != next_type || bam_aux2i(p) != next_val) {
fail("after updating %.2s to %"PRId64":"
" %.2s field is %c:%"PRId64"; expected %c:%"PRId64,
target_id, target_val,
next_id, *p, bam_aux2i(p), next_type, next_val);
return -1;
}
return 0;
}
static int64_t get_mate_score(bam1_t *b) {
uint8_t *data;
int64_t score;
if ((data = bam_aux_get(b, "ms"))) {
score = bam_aux2i(data);
} else {
fprintf(stderr, "[markdup] error: no ms score tag.\n");
return -1;
}
return score;
}
int read_bam(void *data, bam1_t *b)
{
aux_t *aux = (aux_t*)data;
int ret;
while(1) {
uint8_t *tmp = 0;
ret = sam_read1(aux->fp, aux->hdr, b);
if (ret < 0) break;
if (b->core.flag & (BAM_FUNMAP)) continue;
if ((int)b->core.qual < aux->min_mapq) continue;
if (bam_cigar2ulen(b->core.n_cigar, bam_get_cigar(b)) < aux->min_len) continue;
tmp = bam_aux_get(b, "AS");
if (tmp && bam_aux2i(tmp) < aux->min_as) continue;
break;
}
return ret;
}
int main(int argc, char *argv[])
{
short out2stdout=0;
hashtable ht=new_hashtable(HASHSIZE);
bamFile in,in2;
bamFile out;
int paired;//1 if not paired or pair read 1, 2 otherwise
index_mem=sizeof(hashtable)*sizeof(hashnode**)*HASHSIZE*2;
if (argc != 3) {
fprintf(stderr, "Usage: bam_fix_NH <in.bam> <out.bam or - for stdout>\n");
return 1;
}
// Open file and exit if error
in = bam_open(argv[1], "rb");
out2stdout = strcmp(argv[2], "-")? 0 : 1;
out = strcmp(argv[2], "-")? bam_open(argv[2], "w") : bam_dopen(fileno(stdout), "w");
if (in == 0 ) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[1]);
return 1;
}
if (out == 0) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[2]);
return 1;
}
unsigned long num_alns=0;
int ref;
// ***********
// Copy header
bam_header_t *header;
header = bam_header_read(in);
bam_header_write(out,header);
// sorted by name?
// Should not rely on the value in SO
bam1_t *aln=bam_init1();
bam1_t *prev=bam_init1();
if (!out2stdout) {
fprintf(stderr,"bam_fix_NH version %s\n",VERSION);
fprintf(stderr,"Processing %s\n",argv[1]);
fprintf(stderr,"Hashing...\n");fflush(stderr);
}
while(bam_read1(in,aln)>=0) { // read alignment
if (aln->core.tid < 0) continue;//ignore unaligned reads
if (aln->core.flag & BAM_FUNMAP) continue;
if (aln->core.flag & BAM_FREAD2) paired=2;
else paired=1;
++num_alns;
new_read_aln(ht,fix_read_name(bam1_qname(aln),paired));
if(!out2stdout) PRINT_ALNS_PROCESSED(num_alns);
}
bam_close(in);
if(!out2stdout) {
fprintf(stderr,"%s%lu\n",BACKLINE,num_alns);
fprintf(stderr,"Hashing complete (%lu alignments)\n",num_alns);
fprintf(stderr,"Memory used: %ld MB\n",index_mem/1024/1024);
fprintf(stderr,"Updating entries with NH and printing BAM...\n");
fflush(stderr);
}
// reopen
in2 = bam_open(argv[1], "rb");
if (in2 == 0 ) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[1]);
return 1;
}
header = bam_header_read(in2);
num_alns=0;
while(bam_read1(in2,aln)>=0) { // read alignment
paired=1;
if (aln->core.tid < 0) continue;//ignore unaligned reads
if (aln->core.flag & BAM_FUNMAP) continue;
if (aln->core.flag & BAM_FREAD2) paired=2;
++num_alns;
READ_ALN *r=get_read_aln(ht,fix_read_name(bam1_qname(aln),paired));
assert(r!=NULL);
// update the NH field
uint8_t *old_nh = bam_aux_get(aln, "NH");
int32_t nh=r->ctr;
if (old_nh) {
if (nh!=bam_aux2i(old_nh)) {
fprintf(stderr,"warning: value mismatch! replacing>%s %d->%d\n",bam1_qname(aln),bam_aux2i(old_nh),nh);
}
bam_aux_del(aln, old_nh);
bam_aux_append(aln, "NH", 'i', 4, (uint8_t*)&nh);
#ifdef DEBUG
// printf("!>%s %d\n",bam1_qname(aln),r->ctr);
#endif
}
if (!old_nh) { // add NH
bam_aux_append(aln, "NH", 'i', 4, (uint8_t*)&nh);
#ifdef DEBUG
fprintf(stderr,"!>%s %d\n",bam1_qname(aln),bam_aux2i(old_nh));
#endif
}
bam_write1(out,aln);
if(!out2stdout) PRINT_ALNS_PROCESSED(num_alns);
}
//
bam_destroy1(aln);
bam_close(in2);
bam_close(out);
if(!out2stdout) {
fprintf(stderr,"%s%lu\n",BACKLINE,num_alns);
fprintf(stderr,"Done.\n");
}
return 0;
}
void signalFromBAM(const string bamFileName, const string sigFileName, Parameters P) {
bam1_t *bamA;
bamA=bam_init1();
double nMult=0, nUniq=0;
if (P.outWigFlags.norm==1) {//count reads in the BAM file
BGZF *bamIn=bgzf_open(bamFileName.c_str(),"r");
bam_hdr_t *bamHeader=bam_hdr_read(bamIn);
while ( true ) {//until the end of file
int bamBytes1=bam_read1(bamIn, bamA);
if (bamBytes1<0) break; //end of file
if (bamA->core.tid<0) continue; //unmapped read
// if ( !std::regex_match(chrName.at(bamA->core.tid),std::regex(P.outWigReferencesPrefix))) continue; //reference does not mathc required references
if ( P.outWigReferencesPrefix!="-" && (P.outWigReferencesPrefix.compare(0,P.outWigReferencesPrefix.size(),bamHeader->target_name[bamA->core.tid],P.outWigReferencesPrefix.size())!=0) ) continue; //reference does not match required references
uint8_t* aNHp=bam_aux_get(bamA,"NH");
if (aNHp!=NULL) {
uint32_t aNH=bam_aux2i(aNHp);
if (aNH==1) {//unique mappers
++nUniq;
} else if (aNH>1) {
nMult+=1.0/aNH;
};
};
};
bgzf_close(bamIn);
};
BGZF *bamIn=bgzf_open(bamFileName.c_str(),"r");
bam_hdr_t *bamHeader=bam_hdr_read(bamIn);
int sigN=P.outWigFlags.strand ? 4 : 2;
double *normFactor=new double[sigN];
ofstream **sigOutAll=new ofstream* [sigN];
string* sigOutFileName=new string[sigN];
sigOutFileName[0]=sigFileName+".Unique.str1.out";
sigOutFileName[1]=sigFileName+".UniqueMultiple.str1.out";
if (P.outWigFlags.strand) {
sigOutFileName[2]=sigFileName+".Unique.str2.out";
sigOutFileName[3]=sigFileName+".UniqueMultiple.str2.out";
};
for (int ii=0; ii<sigN; ii++) {
sigOutFileName[ii]+= (P.outWigFlags.format==0 ? ".bg" : ".wig");
sigOutAll[ii]=new ofstream ( sigOutFileName[ii].c_str() );
};
if (P.outWigFlags.norm==0) {//raw counts
normFactor[0]=1;
normFactor[1]=1;
} else if (P.outWigFlags.norm==1) {//normlaized
normFactor[0]=1.0e6 / nUniq;
normFactor[1]=1.0e6 / (nUniq+nMult);
for (int is=0;is<sigN;is++) {//formatting double output
*sigOutAll[is]<<setiosflags(ios::fixed) << setprecision(5);
};
};
if (P.outWigFlags.strand) {
normFactor[2]=normFactor[0];
normFactor[3]=normFactor[1];
};
int iChr=-999;
double *sigAll=NULL;
uint32_t chrLen=0;
while ( true ) {//until the end of file
int bamBytes1=bam_read1(bamIn, bamA);
if (bamA->core.tid!=iChr || bamBytes1<0) {
//output to file
if (iChr!=-999) {//iChr=-999 marks chromosomes that are not output, including unmapped reads
for (int is=0;is<sigN;is++) {
if (P.outWigFlags.format==1) {
*sigOutAll[is] <<"variableStep chrom="<<bamHeader->target_name[iChr] <<"\n";
};
double prevSig=0;
for (uint32_t ig=0;ig<chrLen;ig++) {
double newSig=sigAll[sigN*ig+is];
if (P.outWigFlags.format==0) {//bedGraph
if (newSig!=prevSig) {
if (prevSig!=0) {//finish previous record
*sigOutAll[is] <<ig<<"\t"<<prevSig*normFactor[is] <<"\n"; //1-based end
};
if (newSig!=0) {
*sigOutAll[is] << bamHeader->target_name[iChr] <<"\t"<< ig <<"\t"; //0-based beginning
};
prevSig=newSig;
};
} else if (P.outWigFlags.format==1){//wiggle
if (newSig!=0) {
*sigOutAll[is] <<ig+1<<"\t"<<newSig*normFactor[is] <<"\n";
};
};
};
};
};
if (bamBytes1<0) {//no more reads
break;
};
iChr=bamA->core.tid;
if ( iChr==-1 || (P.outWigReferencesPrefix!="-" && (P.outWigReferencesPrefix.compare(0,P.outWigReferencesPrefix.size(),bamHeader->target_name[bamA->core.tid],P.outWigReferencesPrefix.size())!=0) ) ) {
iChr=-999;
continue; //reference does not match required references
};
chrLen=bamHeader->target_len[iChr]+1;//one extra base at the end which sohuld always be 0
delete [] sigAll;
sigAll= new double[sigN*chrLen];
memset(sigAll, 0, sizeof(*sigAll)*sigN*chrLen);
};
// uint32_t nCigar =(bamA->core.flag<<16)>>16;
// uint32_t mapFlag=bamA->core.flag>>16;
// uint32_t mapQ=(bamA->core.flag<<16)>>24;
#define BAM_CIGAR_OperationShift 4
#define BAM_CIGAR_LengthBits 28
#define BAM_CIGAR_M 0
#define BAM_CIGAR_I 1
#define BAM_CIGAR_D 2
#define BAM_CIGAR_N 3
#define BAM_CIGAR_S 4
#define BAM_CIGAR_H 5
#define BAM_CIGAR_P 6
#define BAM_CIGAR_EQ 7
#define BAM_CIGAR_X 8
//by default, alignments marked as duplicate are not processed
if ( (bamA->core.flag & 0x400) > 0 ) continue;
//NH attribute
uint8_t* aNHp=bam_aux_get(bamA,"NH");
uint32_t aNH;
if (aNHp==NULL) {
aNH=1; //no NH tag: assume NH=1
//continue; //do not process lines without NH field
} else {
aNH=bam_aux2i(bam_aux_get(bamA,"NH")); //write a safer function allowing for lacking NH tag
};
if (aNH==0) continue; //do not process lines without NH=0
uint32_t aG=bamA->core.pos;
uint32_t iStrand=0;
if (P.outWigFlags.strand) {//strand for stranded data from SAM flag
iStrand= ( (bamA->core.flag & 0x10) > 0 ) == ( (bamA->core.flag & 0x80) == 0 );//0/1 for +/-
};
if (P.outWigFlags.type==1) {//5' of the1st read signal only, RAMPAGE/CAGE
if ( (bamA->core.flag & 0x80)>0) continue; //skip if this the second mate
if (iStrand==0) {
if (aNH==1) {//unique mappers
sigAll[aG*sigN+0+2*iStrand]++;
};
sigAll[aG*sigN+1+2*iStrand]+=1.0/aNH;//U+M, normalized by the number of multi-mapping loci
continue; //record only the first position
};
};
uint32_t* cigar=(uint32_t*) (bamA->data+bamA->core.l_qname);
for (uint32_t ic=0; ic<bamA->core.n_cigar; ic++) {
uint32_t cigOp=(cigar[ic]<<BAM_CIGAR_LengthBits)>>BAM_CIGAR_LengthBits;
uint32_t cigL=cigar[ic]>>BAM_CIGAR_OperationShift;
switch (cigOp) {
case(BAM_CIGAR_D):
case(BAM_CIGAR_N):
aG+=cigL;
break;
case(BAM_CIGAR_M):
if (P.outWigFlags.type==0 || (P.outWigFlags.type==2 && (bamA->core.flag & 0x80)>0 )) {//full signal, or second mate onyl signal
for (uint32_t ig=0;ig<cigL;ig++) {
if (aG>=chrLen) {
cerr << "BUG: alignment extends past chromosome in signalFromBAM.cpp\n";
exit(-1);
};
if (aNH==1) {//unique mappers
sigAll[aG*sigN+0+2*iStrand]++;
};
sigAll[aG*sigN+1+2*iStrand]+=1.0/aNH;//U+M, normalized by the number of multi-mapping loci
aG++;
};
} else {
aG+=cigL;
};
};
};
if (P.outWigFlags.type==1) {//full signal
--aG;
if (aNH==1) {//unique mappers
sigAll[aG*sigN+0+2*iStrand]++;
};
sigAll[aG*sigN+1+2*iStrand]+=1.0/aNH;//U+M, normalized by the number of multi-mapping loci
};
};
delete [] sigAll;
for (int is=0; is<sigN; is++) {// flush/close all signal files
sigOutAll[is]->flush();
sigOutAll[is]->close();
};
};
int main(int argc, char *argv[])
{
bamFile in;
sqlite3 * db;
sqlite3_stmt * stmt;
char * sErrMsg = NULL;
char * tail = 0;
int nRetCode;
char sSQL [BUFFER_SIZE] = "\0";
char database[BUFFER_SIZE];
clock_t startClock,startClock2;
if (argc != 2) {
fprintf(stderr, "Usage: bamRindex <in.bam>\n");
return 1;
}
// Open file and exit if error
//in = strcmp(argv[1], "-")? bam_open(argv[1], "rb") : bam_dopen(fileno(stdin), "rb");
//fprintf(stderr,"Options ok\n");
in = bam_open(argv[1], "rb");
if (in == 0 ) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[1]);
return 1;
}
//fprintf(stderr,"BAM opened\n");
assert(strcpy(database,argv[1])!=NULL);
assert(strcat(database,".ridx")!=NULL);
remove(database);
// ***********
// Read header
bam_header_t *header;
header = bam_header_read(in);
// sorted by name?
// Should not rely on the value in SO
bam1_t *aln=bam_init1();
unsigned long num_alns=0;
/*********************************************/
/* Open the Database and create the Schema */
// TODO: check the errors
sqlite3_open(database, &db);
sqlite3_exec(db, TABLE, NULL, NULL, &sErrMsg); // create the table
SQLITE_CHECK_ERROR();
startClock = clock();
sqlite3_exec(db, "PRAGMA synchronous = 0;", NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
sqlite3_exec(db, "PRAGMA journal_mode = OFF;", NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
// Use up to 8GB of memory
sqlite3_exec(db, "PRAGMA cache_size = -8000000;", NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
sqlite3_exec(db, "BEGIN TRANSACTION;", NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
while(bam_read1(in,aln)>=0) { // read alignment
//aln->core.tid < 0 ?
uint8_t *nh = bam_aux_get(aln, "NH");
uint8_t *nm = bam_aux_get(aln, "NM");
uint8_t *xs = bam_aux_get(aln, "XS");
BOOLEAN isPrimary;
BOOLEAN isMapped;
BOOLEAN notMapped;
BOOLEAN isDuplicate;
BOOLEAN isNotPassingQualityControls;
BOOLEAN isPaired;
BOOLEAN isSecondMateRead,isProperPair;
//secondary alignment
notMapped=(aln->core.flag & BAM_FUNMAP) ? TRUE: FALSE;
//notMapped=((aln->core.flag & BAM_FUNMAP) || (aln->core.mtid ==0)) ? TRUE: FALSE;
isMapped=!notMapped;
isPrimary= (aln->core.flag & BAM_FSECONDARY) ? FALSE:TRUE;
isProperPair=(aln->core.flag & BAM_FPROPER_PAIR) ? TRUE:FALSE;
isPaired=(aln->core.flag & BAM_FPAIRED ) ? TRUE:FALSE;
isSecondMateRead=(aln->core.flag & BAM_FREAD2 ) ? TRUE: FALSE;
isNotPassingQualityControls=(aln->core.flag & BAM_FQCFAIL ) ? TRUE:FALSE;
isDuplicate=(aln->core.flag & BAM_FDUP) ? TRUE: FALSE;
BOOLEAN isSpliced=FALSE;
BOOLEAN hasSimpleCigar=TRUE;
int nSpliced=0;
int i;
if (aln->core.n_cigar != 0) {
for (i = 0; i < aln->core.n_cigar; ++i) {
char l="MIDNSHP=X"[bam1_cigar(aln)[i]&BAM_CIGAR_MASK];
//fprintf(stderr,"%c",l);
if ( l == 'N' ) { isSpliced=TRUE; hasSimpleCigar=FALSE;++nSpliced;}
if ( l != 'M' && l!='=' ) { hasSimpleCigar=FALSE;}
}
}
//fprintf(stderr,"read %ld\n",num_alns);
// isDuplicate,isNotPassingQualityControls,
// isSpliced,isPAired,isPrimary,hasSimpleCigar,isSecondMateRead,isProperPair,nh,nm,qual/mapq,xs
sprintf(sSQL,"INSERT into bam_index values (%d,%d,%d,%d,%d,%d,%d,%d,%d,%d,%d,%d,'%c')",
isDuplicate,isNotPassingQualityControls,
nSpliced,isPaired,isPrimary,isMapped,hasSimpleCigar,isSecondMateRead,isProperPair,
(nh==0?0:bam_aux2i(nh)),(nm==0?0:bam_aux2i(nm)),
aln->core.qual,
(xs==0?' ':(bam_aux2A(xs)==0?' ':bam_aux2A(xs))));
sqlite3_exec(db, sSQL, NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
++num_alns;
PRINT_ALNS_PROCESSED(num_alns);
}
bam_close(in);
sqlite3_exec(db, "END TRANSACTION;", NULL, NULL, &sErrMsg);
SQLITE_CHECK_ERROR();
printf("\nImported %d records in %4.2f seconds\n", num_alns, ( (double) (clock() - startClock))/CLOCKS_PER_SEC);
// Create the indexes
startClock2 = clock();
// generating the indexes does not pay off
//sqlite3_exec(db, INDEXES, NULL, NULL, &sErrMsg);
//printf("Indexed %d records in %4.2f seconds\n", num_alns, ( (double) (clock() - startClock2))/CLOCKS_PER_SEC);
printf("Total time: %4.2f seconds\n", ((double)(clock() - startClock))/CLOCKS_PER_SEC);
sqlite3_close(db);
return 0;
}
void bam_fillmd1_core(bam1_t *b, char *ref, int ref_len, int flag, int max_nm)
{
uint8_t *seq = bam_get_seq(b);
uint32_t *cigar = bam_get_cigar(b);
bam1_core_t *c = &b->core;
int i, x, y, u = 0;
kstring_t *str;
int32_t old_nm_i = -1, nm = 0;
str = (kstring_t*)calloc(1, sizeof(kstring_t));
for (i = y = 0, x = c->pos; i < c->n_cigar; ++i) {
int j, l = cigar[i]>>4, op = cigar[i]&0xf;
if (op == BAM_CMATCH || op == BAM_CEQUAL || op == BAM_CDIFF) {
for (j = 0; j < l; ++j) {
int c1, c2, z = y + j;
if (x+j >= ref_len || ref[x+j] == '\0') break; // out of bounds
c1 = bam_seqi(seq, z), c2 = seq_nt16_table[(int)ref[x+j]];
if ((c1 == c2 && c1 != 15 && c2 != 15) || c1 == 0) { // a match
if (flag&USE_EQUAL) seq[z/2] &= (z&1)? 0xf0 : 0x0f;
++u;
} else {
kputw(u, str);
kputc(ref[x+j], str);
u = 0;
++nm;
}
}
if (j < l) break;
x += l;
y += l;
} else if (op == BAM_CDEL) {
kputw(u, str);
kputc('^', str);
for (j = 0; j < l; ++j) {
if (x+j >= ref_len || ref[x+j] == '\0') break;
kputc(ref[x+j], str);
}
u = 0;
x += j;
nm += j;
if (j < l) break;
} else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) {
y += l;
if (op == BAM_CINS) nm += l;
} else if (op == BAM_CREF_SKIP) {
x += l;
}
}
kputw(u, str);
// apply max_nm
if (max_nm > 0 && nm >= max_nm) {
for (i = y = 0, x = c->pos; i < c->n_cigar; ++i) {
int j, l = cigar[i]>>4, op = cigar[i]&0xf;
if (op == BAM_CMATCH || op == BAM_CEQUAL || op == BAM_CDIFF) {
for (j = 0; j < l; ++j) {
int c1, c2, z = y + j;
if (x+j >= ref_len || ref[x+j] == '\0') break; // out of bounds
c1 = bam_seqi(seq, z), c2 = seq_nt16_table[(int)ref[x+j]];
if ((c1 == c2 && c1 != 15 && c2 != 15) || c1 == 0) { // a match
seq[z/2] |= (z&1)? 0x0f : 0xf0;
bam_get_qual(b)[z] = 0;
}
}
if (j < l) break;
x += l;
y += l;
} else if (op == BAM_CDEL || op == BAM_CREF_SKIP) x += l;
else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) y += l;
}
}
// update NM
if ((flag & UPDATE_NM) && !(c->flag & BAM_FUNMAP)) {
uint8_t *old_nm = bam_aux_get(b, "NM");
if (old_nm) old_nm_i = bam_aux2i(old_nm);
if (!old_nm) bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm);
else if (nm != old_nm_i) {
fprintf(stderr, "[bam_fillmd1] different NM for read '%s': %d -> %d\n", bam_get_qname(b), old_nm_i, nm);
bam_aux_del(b, old_nm);
bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm);
}
}
// update MD
if ((flag & UPDATE_MD) && !(c->flag & BAM_FUNMAP)) {
uint8_t *old_md = bam_aux_get(b, "MD");
if (!old_md) bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s);
else {
int is_diff = 0;
if (strlen((char*)old_md+1) == str->l) {
for (i = 0; i < str->l; ++i)
if (toupper(old_md[i+1]) != toupper(str->s[i]))
break;
if (i < str->l) is_diff = 1;
} else is_diff = 1;
if (is_diff) {
fprintf(stderr, "[bam_fillmd1] different MD for read '%s': '%s' -> '%s'\n", bam_get_qname(b), old_md+1, str->s);
bam_aux_del(b, old_md);
bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s);
}
}
}
// drop all tags but RG
if (flag&DROP_TAG) {
uint8_t *q = bam_aux_get(b, "RG");
bam_aux_drop_other(b, q);
}
// reduce the resolution of base quality
if (flag&BIN_QUAL) {
uint8_t *qual = bam_get_qual(b);
for (i = 0; i < b->core.l_qseq; ++i)
if (qual[i] >= 3) qual[i] = qual[i]/10*10 + 7;
}
free(str->s);
free(str);
}
static int aux_fields1(void)
{
static const char sam[] = "data:,"
"@SQ\tSN:one\tLN:1000\n"
"@SQ\tSN:two\tLN:500\n"
"r1\t0\tone\t500\t20\t8M\t*\t0\t0\tATGCATGC\tqqqqqqqq\tXA:A:k\tXi:i:37\tXf:f:" xstr(PI) "\tXd:d:" xstr(E) "\tXZ:Z:" HELLO "\tXH:H:" BEEF "\tXB:B:c,-2,0,+2\tB0:B:i,-2147483648,-1,0,1,2147483647\tB1:B:I,0,1,2147483648,4294967295\tB2:B:s,-32768,-1,0,1,32767\tB3:B:S,0,1,32768,65535\tB4:B:c,-128,-1,0,1,127\tB5:B:C,0,1,127,255\tBf:B:f,-3.14159,2.71828\tZZ:i:1000000\tF2:d:2.46801\tY1:i:-2147483648\tY2:i:-2147483647\tY3:i:-1\tY4:i:0\tY5:i:1\tY6:i:2147483647\tY7:i:2147483648\tY8:i:4294967295\n";
// Canonical form of the alignment record above, as output by sam_format1()
static const char r1[] = "r1\t0\tone\t500\t20\t8M\t*\t0\t0\tATGCATGC\tqqqqqqqq\tXi:i:37\tXf:f:3.14159\tXd:d:2.71828\tXZ:Z:" NEW_HELLO "\tXH:H:" BEEF "\tXB:B:c,-2,0,2\tB0:B:i,-2147483648,-1,0,1,2147483647\tB1:B:I,0,1,2147483648,4294967295\tB2:B:s,-32768,-1,0,1,32767\tB3:B:S,0,1,32768,65535\tB4:B:c,-128,-1,0,1,127\tB5:B:C,0,1,127,255\tBf:B:f,-3.14159,2.71828\tZZ:i:1000000\tF2:f:9.8765\tY1:i:-2147483648\tY2:i:-2147483647\tY3:i:-1\tY4:i:0\tY5:i:1\tY6:i:2147483647\tY7:i:2147483648\tY8:i:4294967295\tN0:i:-1234\tN1:i:1234\tN2:i:-2\tN3:i:3\tF1:f:4.5678\tN4:B:S,65535,32768,1,0\tN5:i:4242";
samFile *in = sam_open(sam, "r");
bam_hdr_t *header = sam_hdr_read(in);
bam1_t *aln = bam_init1();
uint8_t *p;
kstring_t ks = { 0, 0, NULL };
int64_t b0vals[5] = { -2147483648LL,-1,0,1,2147483647LL }; // i
int64_t b1vals[4] = { 0,1,2147483648LL,4294967295LL }; // I
int64_t b2vals[5] = { -32768,-1,0,1,32767 }; // s
int64_t b3vals[4] = { 0,1,32768,65535 }; // S
int64_t b4vals[5] = { -128,-1,0,1,127 }; // c
int64_t b5vals[4] = { 0,1,127,255 }; // C
// NB: Floats not doubles below!
// See https://randomascii.wordpress.com/2012/06/26/doubles-are-not-floats-so-dont-compare-them/
float bfvals[2] = { -3.14159f, 2.71828f };
int8_t n4v1[] = { -128, -64, -32, -16, -8, -4, -2, -1,
0, 1, 2, 4, 8, 16, 32, 64, 127 };
uint32_t n4v2[] = { 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 1234, 5678, 1U << 31, 0 };
int16_t n4v3[] = { -32768, -1, 0, 1, 32767 };
float n4v4[] = { 0, 1, 2, 10, 20, 30, 1.5, -2.5 };
uint8_t n4v5[] = { 0, 255 };
int32_t n4v6[] = { -2147483647 - 1, 10, -1, 0, 1, 2147483647 };
uint16_t n4v7[] = { 65535, 32768, 1, 0 };
int32_t ival = -1234;
uint32_t uval = 1234;
float f1 = 4.5678;
float f2 = 9.8765;
size_t nvals, i;
if (sam_read1(in, header, aln) >= 0) {
if ((p = check_bam_aux_get(aln, "XA", 'A')) && bam_aux2A(p) != 'k')
fail("XA field is '%c', expected 'k'", bam_aux2A(p));
bam_aux_del(aln,p);
if (bam_aux_get(aln,"XA"))
fail("XA field was not deleted");
if ((p = check_bam_aux_get(aln, "Xi", 'C')) && bam_aux2i(p) != 37)
fail("Xi field is %"PRId64", expected 37", bam_aux2i(p));
if ((p = check_bam_aux_get(aln, "Xf", 'f')) && fabs(bam_aux2f(p) - PI) > 1E-6)
fail("Xf field is %.12f, expected pi", bam_aux2f(p));
if ((p = check_bam_aux_get(aln, "Xd", 'd')) && fabs(bam_aux2f(p) - E) > 1E-6)
fail("Xf field is %.12f, expected e", bam_aux2f(p));
if ((p = check_bam_aux_get(aln, "XZ", 'Z')) && strcmp(bam_aux2Z(p), HELLO) != 0)
fail("XZ field is \"%s\", expected \"%s\"", bam_aux2Z(p), HELLO);
bam_aux_update_str(aln,"XZ",strlen(NEW_HELLO)+1,NEW_HELLO);
if ((p = check_bam_aux_get(aln, "XZ", 'Z')) && strcmp(bam_aux2Z(p), NEW_HELLO) != 0)
fail("XZ field is \"%s\", expected \"%s\"", bam_aux2Z(p), NEW_HELLO);
if ((p = check_bam_aux_get(aln, "XH", 'H')) && strcmp(bam_aux2Z(p), BEEF) != 0)
fail("XH field is \"%s\", expected \"%s\"", bam_aux2Z(p), BEEF);
if ((p = check_bam_aux_get(aln, "XB", 'B'))
&& ! (memcmp(p, "Bc", 2) == 0
&& memcmp(p + 2, "\x03\x00\x00\x00\xfe\x00\x02", 7) == 0))
fail("XB field is %c,..., expected c,-2,0,+2", p[1]);
check_int_B_array(aln, "B0", NELE(b0vals), b0vals);
check_int_B_array(aln, "B1", NELE(b1vals), b1vals);
check_int_B_array(aln, "B2", NELE(b2vals), b2vals);
check_int_B_array(aln, "B3", NELE(b3vals), b3vals);
check_int_B_array(aln, "B4", NELE(b4vals), b4vals);
check_int_B_array(aln, "B5", NELE(b5vals), b5vals);
nvals = NELE(bfvals);
if ((p = check_bam_aux_get(aln, "Bf", 'B')) != NULL) {
if (bam_auxB_len(p) != nvals)
fail("Wrong length reported for Bf field, got %d, expected %zd\n",
bam_auxB_len(p), nvals);
for (i = 0; i < nvals; i++) {
if (bam_auxB2f(p, i) != bfvals[i]) {
fail("Wrong value from bam_auxB2f for Bf field index %zd, "
"got %f expected %f\n",
i, bam_auxB2f(p, i), bfvals[i]);
}
}
}
if ((p = check_bam_aux_get(aln, "ZZ", 'I')) && bam_aux2i(p) != 1000000)
fail("ZZ field is %"PRId64", expected 1000000", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y1")) && bam_aux2i(p) != -2147483647-1)
fail("Y1 field is %"PRId64", expected -2^31", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y2")) && bam_aux2i(p) != -2147483647)
fail("Y2 field is %"PRId64", expected -2^31+1", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y3")) && bam_aux2i(p) != -1)
fail("Y3 field is %"PRId64", expected -1", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y4")) && bam_aux2i(p) != 0)
fail("Y4 field is %"PRId64", expected 0", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y5")) && bam_aux2i(p) != 1)
fail("Y5 field is %"PRId64", expected 1", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y6")) && bam_aux2i(p) != 2147483647)
fail("Y6 field is %"PRId64", expected 2^31-1", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y7")) && bam_aux2i(p) != 2147483648LL)
fail("Y7 field is %"PRId64", expected 2^31", bam_aux2i(p));
if ((p = bam_aux_get(aln, "Y8")) && bam_aux2i(p) != 4294967295LL)
fail("Y8 field is %"PRId64", expected 2^32-1", bam_aux2i(p));
// Try appending some new tags
if (bam_aux_append(aln, "N0", 'i', sizeof(ival), (uint8_t *) &ival) != 0)
fail("Failed to append N0:i tag");
if ((p = bam_aux_get(aln, "N0")) && bam_aux2i(p) != ival)
fail("N0 field is %"PRId64", expected %d", bam_aux2i(p), ival);
if (bam_aux_append(aln, "N1", 'I', sizeof(uval), (uint8_t *) &uval) != 0)
fail("failed to append N1:I tag");
if ((p = bam_aux_get(aln, "N1")) && bam_aux2i(p) != uval)
fail("N1 field is %"PRId64", expected %u", bam_aux2i(p), uval);
// Append tags with bam_aux_update_int()
if (bam_aux_update_int(aln, "N2", -2) < 0)
fail("failed to append N2:c tag");
if (bam_aux_update_int(aln, "N3", 3) < 0)
fail("failed to append N3:C tag");
p = bam_aux_get(aln, "N2");
if (!p)
fail("failed to retrieve N2 tag");
else if (*p != 'c' || bam_aux2i(p) != -2)
fail("N2 field is %c:%"PRId64", expected c:-2", *p, bam_aux2i(p));
p = bam_aux_get(aln, "N3");
if (!p)
fail("failed to retrieve N3 tag");
else if (*p != 'C' || bam_aux2i(p) != 3)
fail("N3 field is %c:%"PRId64", expected C:3", *p, bam_aux2i(p));
// Try changing values with bam_aux_update_int()
i = test_update_int(aln, "N2", 2, 'C', "N3", 3, 'C');
if (i == 0) test_update_int(aln, "N2", 1234, 'S', "N3", 3, 'C');
if (i == 0) test_update_int(aln, "N2", -1, 's', "N3", 3, 'C');
if (i == 0) test_update_int(aln, "N2", 4294967295U, 'I', "N3", 3, 'C');
if (i == 0) test_update_int(aln, "N2", -2, 'i', "N3", 3, 'C');
// Append a value with bam_aux_update_float()
if (bam_aux_update_float(aln, "F1", f1) < 0)
fail("append F1:f tag");
p = bam_aux_get(aln, "F1");
if (!p)
fail("retrieve F1 tag");
else if (*p != 'f' || bam_aux2f(p) != f1)
fail("F1 field is %c:%e, expected f:%e", *p, bam_aux2f(p), f1);
// Change a double tag to a float
if (bam_aux_update_float(aln, "F2", f2) < 0)
fail("update F2 tag");
p = bam_aux_get(aln, "F2");
if (!p)
fail("retrieve F2 tag");
else if (*p != 'f' || bam_aux2f(p) != f2)
fail("F2 field is %c:%e, expected f:%e", *p, bam_aux2f(p), f2);
// Check the next one is intact too
p = bam_aux_get(aln, "Y1");
if (!p)
fail("retrieve Y1 tag");
else if (*p != 'i' && bam_aux2i(p) != -2147483647-1)
fail("Y1 field is %"PRId64", expected -2^31", bam_aux2i(p));
// bam_aux_update_array tests
// append a new array
i = test_update_array(aln, "N4", 'c', NELE(n4v1), n4v1, "\0\0", 0, 0);
// Add a sentinal to check resizes work
if (i == 0) i = test_update_int(aln, "N5", 4242, 'S', "\0\0", 0, 0);
// alter the array tag a few times
if (i == 0)
i = test_update_array(aln, "N4", 'I', NELE(n4v2), n4v2,
"N5", 4242, 'S');
if (i == 0)
i = test_update_array(aln, "N4", 's', NELE(n4v3), n4v3,
"N5", 4242, 'S');
if (i == 0)
i = test_update_array(aln, "N4", 'f', NELE(n4v4), n4v4,
"N5", 4242, 'S');
if (i == 0)
i = test_update_array(aln, "N4", 'c', NELE(n4v5), n4v5,
"N5", 4242, 'S');
if (i == 0)
i = test_update_array(aln, "N4", 'i', NELE(n4v6), n4v6,
"N5", 4242, 'S');
if (i == 0)
i = test_update_array(aln, "N4", 'S', NELE(n4v7), n4v7,
"N5", 4242, 'S');
if (sam_format1(header, aln, &ks) < 0)
fail("can't format record");
if (strcmp(ks.s, r1) != 0)
fail("record formatted incorrectly: \"%s\"", ks.s);
free(ks.s);
}
else fail("can't read record");
bam_destroy1(aln);
bam_hdr_destroy(header);
sam_close(in);
return 1;
}
// from bam_md.c in SAMtools
// modified not fill in the NM tag, and not to start the reference a c->pos
static void
tmap_sam_md1_core(bam1_t *b, char *ref)
{
uint8_t *seq = bam1_seq(b);
uint32_t *cigar = bam1_cigar(b);
bam1_core_t *c = &b->core;
int i, x, y, u = 0;
kstring_t *str;
uint8_t *old_md, *old_nm;
int32_t old_nm_i=-1, nm=0;
str = (kstring_t*)calloc(1, sizeof(kstring_t));
for (i = y = x = 0; i < c->n_cigar; ++i) {
int j, l = cigar[i]>>4, op = cigar[i]&0xf;
if (op == BAM_CMATCH) {
for (j = 0; j < l; ++j) {
int z = y + j;
int c1 = bam1_seqi(seq, z), c2 = bam_nt16_table[(int)ref[x+j]];
if (ref[x+j] == 0) break; // out of boundary
if ((c1 == c2 && c1 != 15 && c2 != 15) || c1 == 0) { // a match
++u;
} else {
ksprintf(str, "%d", u);
kputc(ref[x+j], str);
u = 0;
nm++;
}
}
if (j < l) break;
x += l; y += l;
} else if (op == BAM_CDEL) {
ksprintf(str, "%d", u);
kputc('^', str);
for (j = 0; j < l; ++j) {
if (ref[x+j] == 0) break;
kputc(ref[x+j], str);
}
u = 0;
if (j < l) break;
x += l;
nm += l;
} else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) {
y += l;
if (op == BAM_CINS) nm += l;
} else if (op == BAM_CREF_SKIP) {
x += l;
}
}
ksprintf(str, "%d", u);
// update MD
old_md = bam_aux_get(b, "MD");
if(NULL == old_md) {
bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s);
}
else {
int is_diff = 0;
if(strlen((char*)old_md+1) == str->l) {
for(i = 0; i < str->l; ++i) {
if(toupper(old_md[i+1]) != toupper(str->s[i])) {
break;
}
}
if(i < str->l) {
is_diff = 1;
}
}
else {
is_diff = 1;
}
if(1 == is_diff) {
bam_aux_del(b, old_md);
bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s);
}
}
// update NM
old_nm = bam_aux_get(b, "NM");
if(NULL != old_nm) {
old_nm_i = bam_aux2i(old_nm);
if(old_nm_i != nm) {
bam_aux_del(b, old_nm);
bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm);
}
}
free(str->s); free(str);
}
int main(int argc, char *argv[])
{
hashtable ht=new_hashtable(HASHSIZE);
bamFile in,in2;
bamFile out;
if (argc != 3) {
fprintf(stderr, "Usage: bam_fix_NH <in.bam> <out.bam>\n");
return 1;
}
// Open file and exit if error
//in = strcmp(argv[1], "-")? bam_open(argv[1], "rb") : bam_dopen(fileno(stdin), "rb");
in = bam_open(argv[1], "rb");
out = strcmp(argv[2], "-")? bam_open(argv[2], "w") : bam_dopen(fileno(stdout), "w");
if (in == 0 ) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[1]);
return 1;
}
if (out == 0) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[2]);
return 1;
}
unsigned long num_alns=0;
int ref;
// ***********
// Copy header
bam_header_t *header;
header = bam_header_read(in);
bam_header_write(out,header);
// sorted by name?
// Should not rely on the value in SO
bam1_t *aln=bam_init1();
bam1_t *prev=bam_init1();
printf("Hashing...\n");flush(stdout);
while(bam_read1(in,aln)>=0) { // read alignment
if (aln->core.tid < 0) continue;//ignore unaligned reads
++num_alns;
new_read_aln(ht,bam1_qname(aln));
}
bam_close(in);
printf("Hashing complete (%lu alignments)\n",num_alns);
printf("Memory used in the hash: %ld MB\n",index_mem/1024/1024);
flush(stdout);
// reopen
in2 = bam_open(argv[1], "rb");
if (in2 == 0 ) {
fprintf(stderr, "ERROR: Fail to open BAM file %s\n", argv[1]);
return 1;
}
header = bam_header_read(in2);
while(bam_read1(in2,aln)>=0) { // read alignment
if (aln->core.tid < 0) continue;//ignore unaligned reads
++num_alns;
READ_ALN *r=get_read_aln(ht,bam1_qname(aln));
//assert(r!=NULL);
// update the NH field
uint8_t *old_nh = bam_aux_get(aln, "NH");
uint8_t nh=r->ctr;
if (old_nh) {
if (nh!=bam_aux2i(old_nh)) {
fprintf(stderr,"warning: value mismatch! replacing>%s %d->%d\n",bam1_qname(aln),bam_aux2i(old_nh),nh);
}
bam_aux_del(aln, old_nh);
bam_aux_append(aln, "NH", 'i', 4, (uint8_t*)&nh);
}
if (!old_nh) { // add NH
bam_aux_append(aln, "NH", 'i', 4, (uint8_t*)&nh);
#ifdef DEBUG
printf("!>%s %d\n",bam1_qname(aln),bam_aux2i(old_nh));
#endif
}
// in->header
// Also fix the XS:A tag
// BAM_FREAD1
// BAM_FREAD2
// BAM_FREVERSE the read is mapped to the reverse strand
//bam1_cigar(b)
//BAM_CREF_SKIP 3 CIGAR skip on the reference (e.g. spliced alignment)
//BAM_FREVERSE 16 the read is mapped to the reverse strand
if (aln->core.flag & BAM_FSECONDARY) continue; // skip secondary alignments
if (aln->core.flag & ! BAM_FPAIRED) continue; // not paired
if (aln->core.flag & ! BAM_FPROPER_PAIR) continue; // not a proper pair
if (aln->core.flag & ! BAM_FMUNMAP) continue; // the mate is mapped
if (aln->core.flag & BAM_FSECONDARY) continue; // secundary read
if (aln->core.flag & BAM_FREAD2) continue; // only count each pair once
// core.strand == 0 (f/+) 1 r/-
// flag
// bam1_qname(b)
bam_write1(out,aln);
}
//
bam_destroy1(aln);
bam_close(in2);
bam_close(out);
return 0;
/*
uint8_t *old_nm = bam_aux_get(b, "NM");
90 if (c->flag & BAM_FUNMAP) return;
91 if (old_nm) old_nm_i = bam_aux2i(old_nm);
92 if (!old_nm) bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm);
93 else if (nm != old_nm_i) {
94 fprintf(stderr, "[bam_fillmd1] different NM for read '%s': %d -> %d\n", bam1_qname(b), old_nm_i, nm);
95 bam_aux_del(b, old_nm);
96 bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm);
97 }
*/
}
bool ReadAlign::chimericDetection() {
bool chimRecord=false;
//output chains for out-of-STAR chimeric detection
#ifdef OUTPUT_localChains
{
P->inOut->outLocalChains << readName <<"\t"<< Read0[0] <<"\t"<< Read0[1] << "\n";
for (uint iw=0; iw<nW; iw++) {
for (uint itr=0;itr<nWinTr[iw];itr++) {
P->inOut->outLocalChains << trAll[iw][itr]->maxScore<<"\t"<< trAll[iw][itr]->Chr<<"\t"<<trAll[iw][itr]->Str<<"\t"<<trAll[iw][itr]->nExons;
for (uint ib=0;ib<trAll[iw][itr]->nExons;ib++) {
P->inOut->outLocalChains <<"\t"<< trAll[iw][itr]->exons[ib][EX_G]-P->chrStart[trAll[iw][itr]->Chr] \
<<"\t"<< trAll[iw][itr]->exons[ib][EX_R] <<"\t"<< trAll[iw][itr]->exons[ib][EX_L];
};
P->inOut->outLocalChains <<"\n";
};
};
};
#endif
//////////////////// chimeras
//stich windows => chimeras
//stich only the best window with one of the lower score ones for now - do not stich 2 lower score windows
//stitch only one window on each end of the read
if (P->chimSegmentMin>0 && nW>1 && trBest->rLength >= P->chimSegmentMin \
&& ( trBest->exons[trBest->nExons-1][EX_R] + trBest->exons[trBest->nExons-1][EX_L] + P->chimSegmentMin <= Lread \
|| trBest->exons[0][EX_R] >= P->chimSegmentMin ) \
&& trBest->nextTrScore+P->outFilterMultimapScoreRange < trBest->maxScore \
&& trBest->intronMotifs[0]==0 && (trBest->intronMotifs[1]==0 || trBest->intronMotifs[2]==0) ) {
//there is unmapped space at the start/end, and the main window is not a multimapping window, and non non-canonical junctions, and consistend junction motif
int chimScoreBest=0,chimScoreNext=0;
trChim[0]=*trBest;
uint roStart1=trBest->Str==0 ? trBest->exons[0][EX_R] : Lread - trBest->exons[trBest->nExons-1][EX_R] - trBest->exons[trBest->nExons-1][EX_L];
uint roEnd1=trBest->Str==0 ? trBest->exons[trBest->nExons-1][EX_R] + trBest->exons[trBest->nExons-1][EX_L] - 1 : Lread - trBest->exons[0][EX_R] - 1;
if (roStart1>readLength[0]) roStart1--;
if (roEnd1>readLength[0]) roEnd1--;
uint chimStrBest=0;
if (trBest->intronMotifs[1]==0 && trBest->intronMotifs[2]==0) {//strand is undefined
chimStr=0;
} else if ( (trBest->Str==0) == (trBest->intronMotifs[1]>0)) {//strand the same as RNA
chimStr=1;
} else {//strand opposite to RNA
chimStr=2;
};
for (uint iW=0; iW<nW; iW++) {//check all other windows for chimeras
for (uint iWt=0; iWt<nWinTr[iW]; iWt++){//cycl over transcripts in the window
if (trBest!=trAll[iW][0] && iWt>0) break; //for all windows except that of the best transcript - hceck only iWt=0 (best trnascripts)
if (trBest==trAll[iW][0] && iWt==0) continue;
// {//same window
// if (iWt==0) continue; //do not check the best transcript itself
// if (trBest->exons[0][EX_R]<=trAll[iW][iWt]->exons[0][EX_R]) {
// //start of the last Best exon is before end of the first Chim exon
// if (trBest->exons[trBest->nExons-1][EX_G]<trAll[iW][iWt]->exons[0][EX_G]+trAll[iW][iWt]->exons[0][EX_L]) continue;
// } else {
// if (trAll[iW][iWt]->exons[trAll[iW][iWt]->nExons-1][EX_G]<trBest->exons[0][EX_G]+trBest->exons[0][EX_L]) continue;
// };
// };
if (trAll[iW][iWt]->intronMotifs[0]>0) continue; //do not stitch a window to itself, or to a window with non-canonical junctions
uint chimStr1;
if (trAll[iW][iWt]->intronMotifs[1]==0 && trAll[iW][iWt]->intronMotifs[2]==0) {//strand is undefined
chimStr1=0;
} else if ( (trAll[iW][iWt]->Str==0) == (trAll[iW][iWt]->intronMotifs[1]>0)) {//strand the same as RNA
chimStr1=1;
} else {//strand opposite to RNA
chimStr1=2;
};
if (chimStr!=0 && chimStr1!=0 && chimStr!=chimStr1) continue; //chimeric segments have to have consitent strands
uint roStart2=trAll[iW][iWt]->Str==0 ? trAll[iW][iWt]->exons[0][EX_R] : Lread - trAll[iW][iWt]->exons[trAll[iW][iWt]->nExons-1][EX_R] - trAll[iW][iWt]->exons[trAll[iW][iWt]->nExons-1][EX_L];
uint roEnd2=trAll[iW][iWt]->Str==0 ? trAll[iW][iWt]->exons[trAll[iW][iWt]->nExons-1][EX_R] + trAll[iW][iWt]->exons[trAll[iW][iWt]->nExons-1][EX_L] - 1 : Lread - trAll[iW][iWt]->exons[0][EX_R] - 1;
if (roStart2>readLength[0]) roStart2--;
if (roEnd2>readLength[0]) roEnd2--;
uint chimOverlap = roStart2>roStart1 ? (roStart2>roEnd1 ? 0 : roEnd1-roStart2+1) : (roEnd2<roStart1 ? 0 : roEnd2-roStart1+1);
bool diffMates=(roEnd1 < readLength[0] && roStart2 >= readLength[0]) || (roEnd2 < readLength[0] && roStart1 >= readLength[0]);
//segment lengths && (different mates || small gap between segments)
if (roEnd1 > P->chimSegmentMin + roStart1 + chimOverlap && roEnd2> P->chimSegmentMin + roStart2 + chimOverlap \
&& ( diffMates || ( (roEnd1 + P->maxChimReadGap + 1) >= roStart2 && (roEnd2 + P->maxChimReadGap + 1) >= roStart1 ) ) ) {
//maxChimReadGap=0 in Parameters.cpp
int chimScore=trBest->maxScore + trAll[iW][iWt]->maxScore - (int)chimOverlap; //subtract overlap to avoid double counting
if (chimScore > chimScoreBest && chimScore >= P->chimScoreMin && chimScore+P->chimScoreDropMax >= (int) (readLength[0]+readLength[1]) ) {
trChim[1]=*trAll[iW][iWt];
chimScoreNext=chimScoreBest;
chimScoreBest=chimScore;
trChim[1].roStart = trChim[1].roStr ==0 ? trChim[1].rStart : Lread - trChim[1].rStart - trChim[1].rLength;
trChim[1].cStart = trChim[1].gStart - P->chrStart[trChim[1].Chr];
chimStrBest=chimStr1;
} else if (chimScore>chimScoreNext) {//replace the nextscore if it's not the best one and is higher than the previous one
chimScoreNext=chimScore;
};
};
};//cycle over window transcripts
};//cyecl over windows
if (chimStr==0) chimStr=chimStrBest;
chimN=0;
if (chimScoreNext + P->chimScoreSeparation < chimScoreBest) {//report only if chimera is unique
if (trChim[0].roStart > trChim[1].roStart) swap (trChim[0],trChim[1]);
uint e0 = trChim[0].Str==1 ? 0 : trChim[0].nExons-1;
uint e1 = trChim[1].Str==0 ? 0 : trChim[1].nExons-1;
uint chimRepeat0=0,chimRepeat1=0,chimJ0=0,chimJ1=0;
int chimMotif=0;
chimN=2;
if ( trChim[0].exons[e0][EX_iFrag] > trChim[1].exons[e1][EX_iFrag] ) {//strange configuration, rare, similar to the next one
chimN=0;//reject such chimeras
//good test example:
//CTTAGCTAGCAGCGTCTTCCCAGTGCCTGGAGGGCCAGTGAGAATGGCACCCTCTGGGATTTTTGCTCCTAGGTCT
//TTGAGGTGAAGTTCAAAGATGTGGCTGGCTGTGAGGAGGCCGAGCTAGAGATCATGGAATTTGTGAATTTCTTGAA
} else if ( trChim[0].exons[e0][EX_iFrag] < trChim[1].exons[e1][EX_iFrag] ) {//mates bracket the chimeric junction
chimN=2;
chimRepeat=0;
chimMotif=-1;
if (trChim[0].Str==1) {//negative strand
chimJ0=trChim[0].exons[e0][EX_G]-1;
} else {
chimJ0=trChim[0].exons[e0][EX_G]+trChim[0].exons[e0][EX_L];
};
if (trChim[1].Str==0) {//positive strand
chimJ1=trChim[1].exons[e1][EX_G]-1;
} else {
chimJ1=trChim[1].exons[e1][EX_G]+trChim[1].exons[e1][EX_L];
};
} else {//chimeric junctions is within one of the mates, check and shift chimeric junction if necessary
if (trChim[0].exons[e0][EX_L]>=P->chimJunctionOverhangMin && trChim[1].exons[e1][EX_L]>=P->chimJunctionOverhangMin ) {//large enough overhang required
uint roStart0 = trChim[0].Str==0 ? trChim[0].exons[e0][EX_R] : Lread - trChim[0].exons[e0][EX_R] - trChim[0].exons[e0][EX_L];
uint roStart1 = trChim[1].Str==0 ? trChim[1].exons[e1][EX_R] : Lread - trChim[1].exons[e1][EX_R] - trChim[1].exons[e1][EX_L];
uint jR, jRbest=0;
int jScore=0,jMotif=0,jScoreBest=-999999,jScoreJ=0;
for (jR=0; jR<roStart1+trChim[1].exons[e1][EX_L]-roStart0-1; jR++) {//scan through the exons to find a canonical junction, and check for mismatches
if (jR==readLength[0]) jR++; //skip the inter-mate base
char bR=Read1[0][roStart0+jR];
char b0,b1;
if (trChim[0].Str==0) {
b0=G[trChim[0].exons[e0][EX_G]+jR];
} else {
b0=G[trChim[0].exons[e0][EX_G]+trChim[0].exons[e0][EX_L]-1-jR];
if (b0<4) b0=3-b0;
};
if (trChim[1].Str==0) {
b1=G[trChim[1].exons[e1][EX_G]-roStart1+roStart0+jR];
} else {
b1=G[trChim[1].exons[e1][EX_G]+trChim[1].exons[e1][EX_L]-1+roStart1-roStart0-jR];
if (b1<4) b1=3-b1;
};
if (b0>3 || b1>3 || bR>3) {//chimera is not called if there are Ns in the genome or in the read
chimN=0;
break;
};
char b01,b02,b11,b12;
if (trChim[0].Str==0) {
b01=G[trChim[0].exons[e0][EX_G]+jR+1];
b02=G[trChim[0].exons[e0][EX_G]+jR+2];
} else {
b01=G[trChim[0].exons[e0][EX_G]+trChim[0].exons[e0][EX_L]-1-jR-1];
if (b01<4) b01=3-b01;
b02=G[trChim[0].exons[e0][EX_G]+trChim[0].exons[e0][EX_L]-1-jR-2];
if (b02<4) b02=3-b02;
};
if (trChim[1].Str==0) {
b11=G[trChim[1].exons[e1][EX_G]-roStart1+roStart0+jR-1];
b12=G[trChim[1].exons[e1][EX_G]-roStart1+roStart0+jR];
} else {
b11=G[trChim[1].exons[e1][EX_G]+trChim[1].exons[e1][EX_L]-1+roStart1-roStart0-jR+1];
if (b11<4) b11=3-b11;
b12=G[trChim[1].exons[e1][EX_G]+trChim[1].exons[e1][EX_L]-1+roStart1-roStart0-jR];
if (b12<4) b12=3-b12;
};
jMotif=0;
if (b01==2 && b02==3 && b11==0 && b12==2) {//GTAG
if (chimStr!=2) {
jMotif=1;
};
} else if(b01==1 && b02==3 && b11==0 && b12==1) {//CTAC
if (chimStr!=1) {
jMotif=2;
};
};
if (bR==b0 && bR!=b1) {
jScore++;
} else if (bR!=b0 && bR==b1) {
jScore--;
};
jScoreJ =jMotif==0 ? jScore + P->chimScoreJunctionNonGTAG : jScore ;
if ( jScoreJ > jScoreBest || (jScoreJ == jScoreBest && jMotif>0) ) {
chimMotif=jMotif;
jRbest=jR;
jScoreBest=jScoreJ;
};
};//jR cycle
if (chimN>0) {//else the chimera was rejected because of mismatches
//shift junction in trChim
if (trChim[0].Str==1) {
trChim[0].exons[e0][EX_R] +=trChim[0].exons[e0][EX_L]-jRbest-1;
trChim[0].exons[e0][EX_G] +=trChim[0].exons[e0][EX_L]-jRbest-1;
trChim[0].exons[e0][EX_L]=jRbest+1;
chimJ0=trChim[0].exons[e0][EX_G]-1;
} else {
trChim[0].exons[e0][EX_L]=jRbest+1;
chimJ0=trChim[0].exons[e0][EX_G]+trChim[0].exons[e0][EX_L];
};
if (trChim[1].Str==0) {
trChim[1].exons[e1][EX_R] +=roStart0+jRbest+1-roStart1;
trChim[1].exons[e1][EX_G] +=roStart0+jRbest+1-roStart1;
trChim[1].exons[e1][EX_L]=roStart1+trChim[1].exons[e1][EX_L]-roStart0-jRbest-1;
chimJ1=trChim[1].exons[e1][EX_G]-1;
} else {
trChim[1].exons[e1][EX_L]=roStart1+trChim[1].exons[e1][EX_L]-roStart0-jRbest-1;
chimJ1=trChim[1].exons[e1][EX_G]+trChim[1].exons[e1][EX_L];
};
//find repeats
char b0,b1;
uint jR;
for (jR=0;jR<100;jR++) {//forward check
if (trChim[0].Str==0) {
b0=G[chimJ0+jR];
} else {
b0=G[chimJ0-jR];
if (b0<4) b0=3-b0;
};
if (trChim[1].Str==0) {
b1=G[chimJ1+1+jR];
} else {
b1=G[chimJ1-1-jR];
if (b1<4) b1=3-b1;
};
if (b0!=b1) break;
};
chimRepeat1=jR;
for (jR=0;jR<100;jR++) {//reverse check
if (trChim[0].Str==0) {
b0=G[chimJ0-1-jR];
} else {
b0=G[chimJ0+1+jR];
if (b0<4) b0=3-b0;
};
if (trChim[1].Str==0) {
b1=G[chimJ1-jR];
} else {
b1=G[chimJ1+jR];
if (b1<4) b1=3-b1;
};
if (b0!=b1) break;
};
chimRepeat0=jR;
};//chimN>0
};//large enough overhang
};//chimeric junction is within a mate
//debug
// cout << readName <<"\t"<< (trChim[0].Str==0 ? chimJ1-chimJ0 : chimJ0-chimJ1) << "\t"<< (chimMotif>=0 ? P->alignIntronMax : P->alignMatesGapMax)<<"\n";
// cout << chimRepeat0 <<"\t"<<trChim[0].exons[e0][EX_L]<<"\n";
//chimeric alignments output
if ( chimN==2 && trChim[0].exons[e0][EX_L]>=P->chimJunctionOverhangMin+chimRepeat0 \
&& trChim[1].exons[e1][EX_L]>=P->chimJunctionOverhangMin+chimRepeat1 \
&& ( trChim[0].Str!=trChim[1].Str || trChim[0].Chr!=trChim[1].Chr \
|| (trChim[0].Str==0 ? chimJ1-chimJ0+1LLU : chimJ0-chimJ1+1LLU) > (chimMotif>=0 ? P->alignIntronMax : P->alignMatesGapMax) ) )
{//unique chimeras only && minOverhang1
//&& minOverhang2
//&& (diff str || diff chr ||
//|| gap > (alignIntronMax,alignMatesGapMax) ) negative gap = very large # because of uint
chimRecord=true; //chimeric alignment was recorded
//re-calculate the score for chimeric transcripts
trChim[0].alignScore(Read1, G, P);
trChim[1].alignScore(Read1, G, P);
int chimRepresent=-999, chimType=0;
if (trChim[0].exons[0][EX_iFrag]!=trChim[0].exons[trChim[0].nExons-1][EX_iFrag]) {//tr0 has both mates
chimRepresent = 0;
chimType = 1;
trChim[0].primaryFlag=true;//paired portion is primary
trChim[1].primaryFlag=false;
} else if (trChim[1].exons[0][EX_iFrag]!=trChim[1].exons[trChim[1].nExons-1][EX_iFrag]) {//tr1 has both mates
chimRepresent = 1;
chimType = 1;
trChim[1].primaryFlag=true;//paired portion is primary
trChim[0].primaryFlag=false;
} else if (trChim[0].exons[0][EX_iFrag]!=trChim[1].exons[0][EX_iFrag]) {//tr0 and tr1 are single different mates
chimRepresent = -1;
chimType = 2;
trChim[0].primaryFlag=true;
trChim[1].primaryFlag=true;
} else {//two chimeric segments are on the same mate - this can only happen for single-end reads
chimRepresent = (trChim[0].maxScore > trChim[1].maxScore) ? 0 : 1;
chimType = 3;
trChim[chimRepresent].primaryFlag=true;
trChim[1-chimRepresent].primaryFlag=false;
};
if (P->chimOutType=="WithinBAM") {//BAM output
int alignType, bamN=0, bamIsuppl=-1, bamIrepr=-1;
uint bamBytesTotal=0;//estimate of the total size of all bam records, for output buffering
uint mateChr,mateStart;
uint8_t mateStrand;
for (int itr=0;itr<(int)chimN;itr++) {//generate bam for all chimeric pieces
if (chimType==2) {//PE, encompassing
mateChr=trChim[1-itr].Chr;
mateStart=trChim[1-itr].exons[0][EX_G];
mateStrand=(uint8_t) (trChim[1-itr].Str!=trChim[1-itr].exons[0][EX_iFrag]);
alignType=-1;
} else {//spanning chimeric alignment, could be PE or SE
mateChr=-1;mateStart=-1;mateStrand=0;//no need fot mate info unless this is the supplementary alignment
if (chimRepresent==itr) {
alignType=-1; //this is representative part of chimeric alignment, record is as normal; if encompassing chimeric junction, both are recorded as normal
bamIrepr=( (itr%2)==(trChim[itr].Str) ) ? bamN+1 : bamN;//this is the mate that is chimerically split
} else {//"supplementary" chimeric segment
alignType=( (itr%2)==(trChim[itr].Str) ) ? -12 : -11; //right:left chimeric junction
bamIsuppl=bamN;
if (chimType==1) {//PE alignment, need mate info for the suppl
uint iex=0;
for (;iex<trChim[chimRepresent].nExons-1;iex++) {
if (trChim[chimRepresent].exons[iex][EX_iFrag]!=trChim[itr].exons[0][EX_iFrag]) {
break;
};
};
mateChr=trChim[chimRepresent].Chr;
mateStart=trChim[chimRepresent].exons[iex][EX_G];
mateStrand=(uint8_t) (trChim[chimRepresent].Str!=trChim[chimRepresent].exons[iex][EX_iFrag]);
};
};
};
bamN+=alignBAM(trChim[itr], 1, 1, P->chrStart[trChim[itr].Chr], mateChr, mateStart, mateStrand, \
alignType, NULL, P->outSAMattrOrder, outBAMoneAlign+bamN, outBAMoneAlignNbytes+bamN);
bamBytesTotal+=outBAMoneAlignNbytes[0]+outBAMoneAlignNbytes[1];//outBAMoneAlignNbytes[1] = 0 if SE is recorded
};
//write all bam lines
for (int ii=0; ii<bamN; ii++) {//output all pieces
int tagI=-1;
if (ii==bamIrepr) {
tagI=bamIsuppl;
} else if (ii==bamIsuppl) {
tagI=bamIrepr;
};
if (tagI>=0) {
bam1_t *b;
b=bam_init1();
bam_read1_fromArray(outBAMoneAlign[tagI], b);
uint8_t* auxp=bam_aux_get(b,"NM");
uint32_t auxv=bam_aux2i(auxp);
string tagSA1="SAZ"+P->chrName[b->core.tid]+','+to_string((uint)b->core.pos+1) +',' + ( (b->core.flag&0x10)==0 ? '+':'-') + \
',' + bam_cigarString(b) + ',' + to_string((uint)b->core.qual) + ',' + to_string((uint)auxv) + ';' ;
memcpy( (void*) (outBAMoneAlign[ii]+outBAMoneAlignNbytes[ii]), tagSA1.c_str(), tagSA1.size()+1);//copy string including \0 at the end
outBAMoneAlignNbytes[ii]+=tagSA1.size()+1;
* ( (uint32*) outBAMoneAlign[ii] ) = outBAMoneAlignNbytes[ii]-sizeof(uint32);
};
if (P->outBAMunsorted) outBAMunsorted->unsortedOneAlign(outBAMoneAlign[ii], outBAMoneAlignNbytes[ii], ii>0 ? 0 : bamBytesTotal);
if (P->outBAMcoord) outBAMcoord->coordOneAlign(outBAMoneAlign[ii], outBAMoneAlignNbytes[ii], (iReadAll<<32) );
};
};
for (uint iTr=0;iTr<chimN;iTr++) {//write all chimeric pieces to Chimeric.out.sam/junction
if (P->readNmates==2) {
outputTranscriptSAM(trChim[iTr], chimN, iTr, trChim[1-iTr].Chr, trChim[1-iTr].exons[0][EX_G], (int) (trChim[1-iTr].Str!=trChim[1-iTr].exons[0][EX_iFrag]), -1, NULL, &chunkOutChimSAM);
} else {
outputTranscriptSAM(trChim[iTr], chimN, iTr, -1, -1, -1, -1, NULL, &chunkOutChimSAM);
};
};
//junction + SAMp
chunkOutChimJunction << P->chrName[trChim[0].Chr] <<"\t"<< chimJ0 - P->chrStart[trChim[0].Chr]+1 <<"\t"<< (trChim[0].Str==0 ? "+":"-") \
<<"\t"<< P->chrName[trChim[1].Chr] <<"\t"<< chimJ1 - P->chrStart[trChim[1].Chr]+1 <<"\t"<< (trChim[1].Str==0 ? "+":"-") \
<<"\t"<< chimMotif <<"\t"<< chimRepeat0 <<"\t"<< chimRepeat1 <<"\t"<< readName+1 \
<<"\t"<< trChim[0].exons[0][EX_G] - P->chrStart[trChim[0].Chr]+1 <<"\t"<< outputTranscriptCIGARp(trChim[0]) \
<<"\t"<< trChim[1].exons[0][EX_G] - P->chrStart[trChim[1].Chr]+1 <<"\t"<< outputTranscriptCIGARp(trChim[1]) <<"\n"; //<<"\t"<< trChim[0].exons[0][EX_iFrag]+1 --- no need for that, since trChim[0] is always on the first mate
};
};//chimeric score
};//chimeric search
return chimRecord;
};//END
static inline void famstats_fm_loop(famstats_t *s, bam1_t *b, famstats_fm_settings_t *settings)
{
uint8_t *data;
if(b->core.flag & BAM_FREAD2) return; // Silently skip all read 2s since they have the same FM values.
if((b->core.flag & (BAM_FSECONDARY | BAM_FSUPPLEMENTARY))) {
++s->n_flag_fail;
return;
}
if(b->core.qual < settings->minmq) {
++s->n_mq_fail;
return;
}
const int FM(((data = bam_aux_get(b, "FM")) != nullptr ? bam_aux2i(data) : 0));
const int NP(((data = bam_aux_get(b, "NP")) != nullptr ? bam_aux2i(data) : -1));
int RV(((data = bam_aux_get(b, "RV")) != nullptr ? bam_aux2i(data) : -1));
if(UNLIKELY(FM == 0)) LOG_EXIT("Missing required FM tag. Abort!\n");
if(FM < settings->minFM) {
++s->n_fm_fail;
return;
}
if(bam_itag(b, "FP") == 0) {
++s->n_fp_fail;
if(settings->skip_fp_fail) return;
}
++s->n_pass;
if(FM > 1) {
++s->realfm_counts;
s->realfm_sum += FM;
s->realrc_sum += RV < 0 ? 0 : RV;
}
++s->allfm_counts;
s->allfm_sum += FM;
s->allrc_sum += RV < 0 ? 0 : RV;
int khr;
// Have we seen this family size before?
if((s->ki = kh_get(fm, s->fm, FM)) == kh_end(s->fm))
// If not, put it into the hash table with a count of 1.
s->ki = kh_put(fm, s->fm, FM, &khr), kh_val(s->fm, s->ki) = 1;
else ++kh_val(s->fm, s->ki); // Otherwise increment counts
// Same, but for RV
if((s->ki = kh_get(fm, s->rc, RV)) == kh_end(s->rc))
s->ki = kh_put(fm, s->rc, RV, &khr), kh_val(s->rc, s->ki) = 1;
else ++kh_val(s->rc, s->ki);
// Same, but for NP
if(NP > 0) {
if((s->ki = kh_get(fm, s->np, NP)) == kh_end(s->np))
s->ki = kh_put(fm, s->np, NP, &khr), kh_val(s->np, s->ki) = 1;
else ++kh_val(s->np, s->ki);
}
// If the Duplex Read tag is present, increment duplex read counts
uint8_t *const dr_data(bam_aux_get(b, "DR"));
if(dr_data && bam_aux2i(dr_data)) {
if(RV < 0) RV = 0;
s->dr_sum += FM;
++s->dr_counts;
s->dr_rc_sum += RV;
s->dr_rc_frac_sum += (double)RV / FM;
}
}
bam_stats_t *bam1_stats(bam1_t *bam1, bam_stats_options_t *opts) {
bam_stats_t *bam_stats = NULL;
uint32_t bam_flag = (uint32_t) bam1->core.flag;
if (bam_flag & BAM_FUNMAP) {
// not mapped, then return
bam_stats = bam_stats_new();
bam_stats->mapped = 0;
return bam_stats;
}
if (opts->region_table) {
region_t region;
region.chromosome = opts->sequence_labels[bam1->core.tid];
region.start_position = bam1->core.pos;
region.end_position = region.start_position + bam1->core.l_qseq;
region.strand = NULL;
region.type = NULL;
if (find_region(®ion, opts->region_table)) {
bam_stats = bam_stats_new();
} else {
return NULL;
}
} else {
bam_stats = bam_stats_new();
}
// mapped !!
bam_stats->mapped = 1;
bam_stats->strand = (int) ((bam_flag & BAM_FREVERSE) > 0);
// number of errors
bam_stats->num_errors = bam_aux2i(bam_aux_get(bam1, "NM"));
// cigar handling: number of indels and length
uint32_t cigar_int, *cigar = bam1_cigar(bam1);
int num_cigar_ops = (int) bam1->core.n_cigar;
for (int j = 0; j < num_cigar_ops; j++) {
cigar_int = cigar[j];
switch (cigar_int & BAM_CIGAR_MASK) {
case BAM_CINS: //I: insertion to the reference
case BAM_CDEL: //D: deletion from the reference
bam_stats->num_indels++;
bam_stats->indels_length += (cigar_int >> BAM_CIGAR_SHIFT);
break;
}
}
// quality
bam_stats->quality = bam1->core.qual;
// unique alignment
if (!(bam_flag & BAM_FSECONDARY)) {
bam_stats->unique_alignment = 1;
}
// handling pairs
bam_stats->single_end = 1;
if (bam_flag & BAM_FPAIRED) {
bam_stats->single_end = 0;
if (bam_flag & BAM_FUNMAP) {
if (bam_flag & BAM_FREAD1) {
bam_stats->unmapped_pair_1 = 1;
} else {
bam_stats->unmapped_pair_2 = 1;
}
} else {
if (bam_flag & BAM_FREAD1) {
bam_stats->mapped_pair_1 = 1;
} else {
bam_stats->mapped_pair_2 = 1;
}
}
if (!(bam_flag & BAM_FUNMAP) && !(bam_flag & BAM_FMUNMAP) && (bam_flag & BAM_FPROPER_PAIR)) {
bam_stats->isize = abs(bam1->core.isize);
}
}
// mapping length
char *bam_seq = bam1_seq(bam1);
int seq_len = bam1->core.l_qseq;
bam_stats->seq_length = seq_len;
// nucleotide content
for (int i = 0; i < seq_len; i++) {
switch (bam1_seqi(bam_seq, i)) {
case 1:
bam_stats->num_As++;
break;
case 2:
bam_stats->num_Cs++;
break;
case 4:
bam_stats->num_Gs++;
break;
case 8:
bam_stats->num_Ts++;
break;
case 15:
bam_stats->num_Ns++;
break;
}
}
bam_stats->num_GCs = bam_stats->num_Gs + bam_stats->num_Cs;
return bam_stats;
}
void profileReads(char* bamFile,
int ignoreSuppAlignments,
int ignoreSecondaryAlignments) {
//
int result = -1;
int supp_check = 0x0;
if (ignoreSuppAlignments) {
supp_check |= BAM_FSUPPLEMENTARY;
}
if (ignoreSecondaryAlignments) {
supp_check |= BAM_FSECONDARY;
}
// helper variables
BGZF* in = 0 ;
bam1_t *b = bam_init1();
bam_hdr_t *h;
// open bam
if ((in = bgzf_open(bamFile, "r")) == 0) {
fprintf(stderr,
"ERROR: Failed to open \"%s\" for reading.\n",
bamFile);
}
else if ((h = bam_hdr_read(in)) == 0) { // read header
fprintf(stderr,
"ERROR: Failed to read BAM header of file \"%s\".\n",
bamFile);
}
else {
// destroy header
bam_hdr_destroy(h);
int line = 0;
int supplementary, secondary;
int mapQual;
int matches, mismatches, qLen;
float pcAln, pcId;
int showStats = 0;
uint8_t *aux_mismatches;
// print header
printf("line\tsupp\tsecondary\tmapQ\tmismatches\tmatches\tqLen\tpcId\tpcAln\n");
// fetch alignments
while ((result = bam_read1(in, b)) >= 0) {
line += 1;
// only primary mappings
if ((b->core.flag & supp_check) != 0) {
if (showStats)
fprintf(stdout, "Rejected %d, non-primary\n", line);
continue;
}
supplementary = (b->core.flag & (1 | BAM_FSUPPLEMENTARY)) != 0;
secondary = (b->core.flag & (1 | BAM_FSECONDARY)) != 0;
// quality
mapQual = b->core.qual;
// bam_aux_get returns 0 if optional NM tag is missing
if ((aux_mismatches = bam_aux_get(b, "NM")))
mismatches = bam_aux2i(aux_mismatches);
else
mismatches = 0;
// length
qLen = bam_cigar2qlen((&b->core)->n_cigar, bam_get_cigar(b));
// percent identity
matches = bam_cigar2matches((&b->core)->n_cigar, bam_get_cigar(b));
pcId = (matches - mismatches) / (float)matches; // percentage as float between 0 to 1
// percent alignment
pcAln = matches / (float)qLen; // percentage as float between 0 to 1
// print read values
printf("%d\t%d\t%d\t%d\t%d\t%d\t%d\t%.4f\t%.4f\n",
line, supplementary, secondary, mapQual, mismatches, matches,
qLen, pcId, pcAln);
}
if (result < -1) {
fprintf(stderr,
"ERROR: retrieval of read no. %d from file \"%s\" failed with code %d.\n",
line, bamFile, result);
}
}
if (in) bgzf_close(in);
bam_destroy1(b);
}
static int fetch_func(const bam1_t *b, void *data)
{
fetch_data_t *d = (fetch_data_t*) data;
char *name = bam1_qname(b);
//check if name is requested here
if(!d->requestedTranscripts->empty()) {
//we're doing transcript filtering
if(d->requestedTranscripts->find(name) == d->requestedTranscripts->end()) {
//transcript wasn't requested
return 0;
}
}
fprintf(stderr,"%s\n",name);
//TODO Desparately need some error checking on flag retrieval
char *status = bam_aux2Z(bam_aux_get(b,"YT"));
int length = bam_aux2i(bam_aux_get(b,"HI"));
YTranscript* transcript;
YTranscriptSubStructure structure;
structure.position = b->core.pos + 1;
structure.length = b->core.l_qseq;
structure.ordinal = bam_aux2i(bam_aux_get(b,"HI"));
if(d->transcriptNames.find(name,&transcript)) {
//then we've already found some part of this transcript
transcript->orderedStructures.push_back(structure);
}
else {
transcript = new YTranscript;
char *tidName = d->in->header->target_name[b->core.tid];
char *refName = new char[ strlen(tidName) + 1];
strcpy(refName, tidName);
char *transcriptName = new char[ strlen(name) + 1];
strcpy(transcriptName, name);
char *flagGeneName = bam_aux2Z(bam_aux_get(b,"YG"));
char *geneName = new char[ strlen(flagGeneName) + 1];
strcpy(geneName, flagGeneName);
char *statusName = new char[ strlen(status + 1) ];
strcpy(statusName, status);
transcript->gene = geneName;
transcript->name = transcriptName;
transcript->refName = refName;
transcript->status = statusName;
transcript->orderedStructures.push_back(structure);
transcript->strand = b->core.flag & BAM_FREVERSE ? -1 : 1;
transcript->totalNumberOfStructures = bam_aux2i(bam_aux_get(b, "IH"));
transcript->length = length;
d->transcriptNames.insert(name,transcript);
d->transcripts->push_back(transcript);
}
return 0;
}
void filterReads(char * inBamFile,
char * outBamFile,
int minMapQual,
int minLen,
int maxMisMatches,
float minPcId,
float minPcAln,
int ignoreSuppAlignments,
int ignoreSecondaryAlignments) {
//
int result = -1;
int outResult = -1;
int supp_check = 0x0;
if (ignoreSuppAlignments) {
supp_check |= BAM_FSUPPLEMENTARY;
}
if (ignoreSecondaryAlignments) {
supp_check |= BAM_FSECONDARY;
}
// helper variables
BGZF* in = 0;
BGZF* out = 0;
bam1_t *b = bam_init1();
bam_hdr_t *h;
// open bam
if ((in = bgzf_open(inBamFile, "r")) == 0) {
fprintf(stderr,
"ERROR: Failed to open \"%s\" for reading.\n",
inBamFile);
}
else if ((h = bam_hdr_read(in)) == 0) { // read header
fprintf(stderr,
"ERROR: Failed to read BAM header of file \"%s\".\n",
inBamFile);
}
else if ((out = bgzf_open(outBamFile, "w")) == 0) {
fprintf(stderr,
"ERROR: Failed to open \"%s\" for writing.\n",
outBamFile);
}
else {
// write and destroy header
bam_hdr_write(out, h);
bam_hdr_destroy(h);
int line = 0;
int matches, mismatches, qLen;
float pcAln, pcId;
int showStats = 0;
// fetch alignments
while ((result = bam_read1(in, b)) >= 0) {
line += 1;
// only primary mappings
if ((b->core.flag & supp_check) != 0) {
if (showStats)
fprintf(stdout, "Rejected %d, non-primary\n", line);
continue;
}
// only high quality
if (b->core.qual < minMapQual) {
if (showStats)
fprintf(stdout, "Rejected %d, quality: %d\n", line, b->core.qual);
continue;
}
// not too many absolute mismatches
mismatches = bam_aux2i(bam_aux_get(b, "NM"));
if (mismatches > maxMisMatches) {
if (showStats)
fprintf(stdout, "Rejected %d, mismatches: %d\n", line, mismatches);
continue;
}
// not too short
qLen = bam_cigar2qlen((&b->core)->n_cigar, bam_get_cigar(b));
if (qLen < minLen) {
if (showStats)
fprintf(stdout, "Rejected %d, length: %d\n", line, qLen);
continue;
}
// only high percent identity
matches = bam_cigar2matches((&b->core)->n_cigar, bam_get_cigar(b));
pcId = (matches - mismatches) / (float)matches; // percentage as float between 0 to 1
if (pcId < minPcId) {
if (showStats)
fprintf(stdout, "Rejected %d, identity pc: %.4f\n", line, pcId);
continue;
}
// only high percent alignment
pcAln = matches / (float)qLen; // percentage as float between 0 to 1
if (pcAln < minPcAln) {
if (showStats)
fprintf(stdout, "Rejected %d, alignment pc: %.4f\n", line, pcAln);
continue;
}
if ((outResult = bam_write1(out, b)) < -1) {
fprintf(stderr,
"ERROR: Attempt to write read no. %d to file \"%s\" failed with code %d.\n",
line, outBamFile, outResult);
}
}
if (result < -1) {
fprintf(stderr,
"ERROR: retrieval of read no. %d from file \"%s\" failed with code %d.\n",
line, inBamFile, result);
}
}
if (in) bgzf_close(in);
if (out) bgzf_close(out);
bam_destroy1(b);
}
BM_mappedRead * extractReads(char * bamFile,
char ** contigs,
int numContigs,
uint16_t * groups,
char * prettyName,
int headersOnly,
int minMapQual,
int maxMisMatches,
int ignoreSuppAlignments,
int ignoreSecondaryAlignments) {
//-----
// code uses the pattern outlined in samtools view (sam_view.c)
// thanks lh3!
//
int i = 0;
int result = -1;
int hh = 0;
int supp_check = 0x0; // include supp mappings
if (ignoreSuppAlignments) {
supp_check |= BAM_FSUPPLEMENTARY;
}
if (ignoreSecondaryAlignments) {
supp_check |= BAM_FSECONDARY;
}
// we need to let the users know if their pairings
// will be corrupted
int p_corrupt = 0;
// helper variables
samFile *in = 0;
bam_hdr_t *header = NULL;
bam1_t *b = bam_init1();
BM_mappedRead * root = 0;
BM_mappedRead * prev = 0;
// open file handlers
if ((in = sam_open(bamFile, "r")) == 0) {
fprintf(stderr,
"ERROR: Failed to open \"%s\" for reading.\n",
bamFile);
}
else {
// retrieve the header
if ((header = sam_hdr_read(in)) == 0) {
fprintf(stderr,
"ERROR: Failed to read the header from \"%s\".\n",
bamFile);
}
else {
// check the index is intact
hts_idx_t *idx = sam_index_load(in, bamFile); // load index
if (idx == 0) { // index is unavailable
fprintf(stderr,
"ERROR: Random retrieval only works "\
"for indexed files.\n");
}
else {
cfuhash_table_t *pair_buffer = \
cfuhash_new_with_initial_size(1000000);
cfuhash_set_flag(pair_buffer, CFUHASH_FROZEN_UNTIL_GROWS);
for (hh = 0; hh < numContigs; ++hh) {
// parse a region in the format like `chr2:100-200'
hts_itr_t *iter = sam_itr_querys(idx, header, contigs[hh]);
if (iter == NULL) { // reference name is not found
fprintf(stderr,
"WARNING: Could not find contig: "\
"[%s] in BAM: [%s].\n",
contigs[hh],
bamFile);
}
// fetch alignments
int line = 0;
while ((result = sam_itr_next(in, iter, b)) >= 0) {
bam1_core_t core = b->core;
line += 1;
// only high quality?, primary? mappings
if ( core.qual < minMapQual)
continue;
if ((core.flag & supp_check) != 0)
continue;
if(bam_aux2i(bam_aux_get(b, "NM")) > maxMisMatches) {
continue;
}
char * seqId = bam_get_qname(b);
char * seq = 0;
char * qual = 0;
int qual_len = 0;
int seq_len = 0;
// get sequence and quality
if(0 == headersOnly) {
// no point allocating unused space
seq = calloc(core.l_qseq+1, sizeof(char));
qual = calloc(core.l_qseq+1, sizeof(char));
uint8_t *s = bam_get_seq(b);
if (core.flag&BAM_FREVERSE) {
// reverse the read
int r = 0;
for (i = core.l_qseq-1; i >=0 ; --i) {
seq[r]="=TGKCYSBAWRDMHVN"[bam_seqi(s,
i)];
++r;
}
}
else {
for (i = 0; i < core.l_qseq; ++i) {
seq[i]="=ACMGRSVTWYHKDBN"[bam_seqi(s,
i)];
}
}
seq_len = core.l_qseq;
s = bam_get_qual(b);
if (s[0] != 0xff) {
qual_len = core.l_qseq;
for (i = 0; i < core.l_qseq; ++i) {
qual[i] = (char)(s[i] + 33);
}
}
else if (qual != 0) {
free(qual);
qual = 0;
}
}
// work out pairing information
uint8_t rpi = RPI_ERROR;
if (core.flag&BAM_FPAIRED) {
if(core.flag&BAM_FMUNMAP) {
if (core.flag&BAM_FREAD1) {
rpi = RPI_SNGL_FIR;
}
else if (core.flag&BAM_FREAD2) {
rpi = RPI_SNGL_SEC;
}
}
else {
if (core.flag&BAM_FREAD1) {
rpi = RPI_FIR;
}
else if (core.flag&BAM_FREAD2) {
rpi = RPI_SEC;
}
}
}
else {
rpi = RPI_SNGL;
}
// make the funky Id
#define MAX_SEQ_ID_LEN 80
char * seq_id = calloc(MAX_SEQ_ID_LEN,
sizeof(char));
// allocate the string to the buffer but check to
// ensure we're not cutting anything off
int id_len = snprintf(seq_id,
MAX_SEQ_ID_LEN,
"b_%s;c_%s;r_%s",
prettyName,
contigs[hh],
seqId);
if(id_len >= MAX_SEQ_ID_LEN) {
seq_id = calloc(id_len+1, sizeof(char));
snprintf(seq_id,
id_len+1, // don't forget the NULL!
"b_%s;c_%s;r_%s",
prettyName,
contigs[hh],
seqId);
}
// make the mapped read struct
prev = makeMappedRead(seq_id,
seq,
qual,
id_len,
seq_len,
qual_len,
rpi,
groups[hh],
prev);
if (0 == root) { root = prev; }
if(rpi == RPI_SNGL || \
rpi == RPI_SNGL_FIR || \
rpi == RPI_SNGL_SEC) {
// we can just add away
// indicate singleton reads by pointing the
// partner pointer to itself
prev->partnerRead = prev;
}
else {
// RPI_FIR or RPI_SEC
// work out pairing information using the hash
// we append a 1 or 2 to the end so that
// we don't accidentally pair 1's with 1's etc.
char * stripped_result;
if(rpi == RPI_FIR) {
stripped_result = \
pairStripper(seqId,
core.l_qname-1,
'2');
}
else {
stripped_result = \
pairStripper(seqId,
core.l_qname-1,
'1');
}
char * stripped = seqId;
if(stripped_result)
stripped = stripped_result;
//fprintf(stdout, "SEARCH %s\n", stripped);
// now stripped always holds a stripped value
// see if it is in the hash already
BM_mappedRead * stored_MR = \
cfuhash_get(pair_buffer,
stripped);
if (0 != stored_MR) {
// exists in the hash -> Add the pair info
if(rpi == RPI_FIR) {
prev->partnerRead = stored_MR;
}
else {
stored_MR->partnerRead = prev;
}
// delete the entry from the hash
cfuhash_delete(pair_buffer,
stripped);
}
else {
// we should put it in the hash
// make sure to change it into something
// we will find next time
if(rpi == RPI_FIR)
stripped[strlen(stripped)-1] = '1';
else
stripped[strlen(stripped)-1] = '2';
// check to make sure we're not overwriting
// anything important. cfuhash overwrites
// duplicate entries, so we need to grab
// it and put it to "SNGL_XXX" before we
// lose the pointer
BM_mappedRead * OWMMR = \
cfuhash_put(pair_buffer,
stripped, prev);
if(OWMMR) {
if(OWMMR->rpi == RPI_FIR)
OWMMR->rpi = RPI_SNGL_FIR;
else
OWMMR->rpi = RPI_SNGL_SEC;
OWMMR->partnerRead = OWMMR;
printPairCorruptionWarning(p_corrupt);
p_corrupt = 1;
}
}
if(stripped_result != 0) { // free this!
free(stripped_result);
stripped_result = 0;
}
}
}
hts_itr_destroy(iter);
if (result < -1) {
fprintf(stderr, "ERROR: retrieval of reads from "\
"contig: \"%s\" failed due to "\
"truncated file or corrupt BAM index "\
"file\n", header->target_name[hh]);
break;
}
}
// any entries left in the hash are pairs whose mates did
// not meet quality standards
size_t key_size = 0;
char * key;
BM_mappedRead * LOMMR;
size_t pr_size = 1;
if(cfuhash_each_data(pair_buffer,
(void**)&key,
&key_size,
(void**)&LOMMR,
&pr_size)) {
do {
// get the mapped read
// update it's pairing so we know it's really single
if (LOMMR->rpi == RPI_FIR)
LOMMR->rpi = RPI_SNGL_FIR;
else if (LOMMR->rpi == RPI_SEC)
LOMMR->rpi = RPI_SNGL_SEC;
// indicate singleton reads by pointing the
// partner pointer to itself
LOMMR->partnerRead = LOMMR;
} while(cfuhash_next_data(pair_buffer,
(void**)&key,
&key_size,
(void**)&LOMMR,
&pr_size));
}
cfuhash_clear(pair_buffer);
cfuhash_destroy(pair_buffer);
}
hts_idx_destroy(idx); // destroy the BAM index
}
}
// always do this
if (in) sam_close(in);
bam_destroy1(b);
if ( header ) bam_hdr_destroy(header);
return root;
}
void bam_filter(array_list_t *bam1s, array_list_t *passed_bam1s,
array_list_t *failed_bam1s, bam_filter_options_t *opts) {
bam1_t *bam1;
uint32_t bam_flag;
int value;
region_t region;
char **chromosomes;
int num_chromosomes;
if (opts->region_table) {
chromosomes = opts->region_table->ordering;
num_chromosomes = opts->region_table->max_chromosomes;
region.strand = NULL;
region.type = NULL;
}
size_t num_items = array_list_size(bam1s);
for (size_t i = 0; i < num_items; i++) {
bam1 = array_list_get(i, bam1s);
bam_flag = (uint32_t) bam1->core.flag;
// if not mapped,
if (bam_flag & BAM_FUNMAP) {
array_list_insert(bam1, failed_bam1s);
continue;
}
// unique
if (opts->unique && (bam_flag & BAM_FSECONDARY)) {
array_list_insert(bam1, failed_bam1s);
continue;
}
// proper pairs
if (opts->proper_pairs && (bam_flag & BAM_FPAIRED)) {
if ( !(bam_flag & BAM_FPROPER_PAIR)) {
array_list_insert(bam1, failed_bam1s);
continue;
}
}
// length
value = bam1->core.l_qseq;
if (value < opts->min_length || value > opts->max_length) {
array_list_insert(bam1, failed_bam1s);
continue;
}
// quality
value = bam1->core.qual;
if (value < opts->min_quality || value > opts->max_quality) {
array_list_insert(bam1, failed_bam1s);
continue;
}
// num. error
value = bam_aux2i(bam_aux_get(bam1, "NM"));
if (value < opts->min_num_errors || value > opts->max_num_errors) {
array_list_insert(bam1, failed_bam1s);
continue;
}
// region
if (opts->region_table) {
int seq_id = bam1->core.tid;
if (seq_id >=0 && seq_id < num_chromosomes) {
region.chromosome = chromosomes[bam1->core.tid];
region.start_position = bam1->core.pos;
region.end_position = region.start_position + bam1->core.l_qseq;
if (!find_exact_region(®ion, opts->region_table)) {
array_list_insert(bam1, failed_bam1s);
continue;
}
} else {
array_list_insert(bam1, failed_bam1s);
continue;
}
}
// finally, this bam1 passed all the filters,
// insert it in the output list
array_list_insert(bam1, passed_bam1s);
}
}
