/*
* This function calculates ct tag for two bams, it assumes they are from the same template and
* writes the tag to the first read in position terms.
*/
static void bam_template_cigar(bam1_t *b1, bam1_t *b2, kstring_t *str)
{
bam1_t *swap;
int i, end;
uint32_t *cigar;
str->l = 0;
if (b1->core.tid != b2->core.tid || b1->core.tid < 0 || b1->core.pos < 0 || b2->core.pos < 0 || b1->core.flag&BAM_FUNMAP || b2->core.flag&BAM_FUNMAP) return; // coordinateless or not on the same chr; skip
if (b1->core.pos > b2->core.pos) swap = b1, b1 = b2, b2 = swap; // make sure b1 has a smaller coordinate
kputc((b1->core.flag & BAM_FREAD1)? '1' : '2', str); // segment index
kputc((b1->core.flag & BAM_FREVERSE)? 'R' : 'F', str); // strand
for (i = 0, cigar = bam_get_cigar(b1); i < b1->core.n_cigar; ++i) {
kputw(bam_cigar_oplen(cigar[i]), str);
kputc(bam_cigar_opchr(cigar[i]), str);
}
end = bam_endpos(b1);
kputw(b2->core.pos - end, str);
kputc('T', str);
kputc((b2->core.flag & BAM_FREAD1)? '1' : '2', str); // segment index
kputc((b2->core.flag & BAM_FREVERSE)? 'R' : 'F', str); // strand
for (i = 0, cigar = bam_get_cigar(b2); i < b2->core.n_cigar; ++i) {
kputw(bam_cigar_oplen(cigar[i]), str);
kputc(bam_cigar_opchr(cigar[i]), str);
}
uint8_t* data;
if ((data = bam_aux_get(b1,"ct")) != NULL) bam_aux_del(b1, data);
if ((data = bam_aux_get(b2,"ct")) != NULL) bam_aux_del(b2, data);
bam_aux_append(b1, "ct", 'Z', str->l+1, (uint8_t*)str->s);
}
void bam_template_cigar(bam1_t *b1, bam1_t *b2, kstring_t *str)
{
bam1_t *swap;
int i, end;
uint32_t *cigar;
str->l = 0;
if (b1->core.tid != b2->core.tid || b1->core.tid < 0) return; // coordinateless or not on the same chr; skip
if (b1->core.pos > b2->core.pos) swap = b1, b1 = b2, b2 = swap; // make sure b1 has a smaller coordinate
kputc((b1->core.flag & BAM_FREAD1)? '1' : '2', str); // segment index
kputc((b1->core.flag & BAM_FREVERSE)? 'R' : 'F', str); // strand
for (i = 0, cigar = bam1_cigar(b1); i < b1->core.n_cigar; ++i) {
kputw(bam_cigar_oplen(cigar[i]), str);
kputc(bam_cigar_opchr(cigar[i]), str);
}
end = bam_calend(&b1->core, cigar);
kputw(b2->core.pos - end, str);
kputc('T', str);
kputc((b2->core.flag & BAM_FREAD1)? '1' : '2', str); // segment index
kputc((b2->core.flag & BAM_FREVERSE)? 'R' : 'F', str); // strand
for (i = 0, cigar = bam1_cigar(b2); i < b2->core.n_cigar; ++i) {
kputw(bam_cigar_oplen(cigar[i]), str);
kputc(bam_cigar_opchr(cigar[i]), str);
}
bam_aux_append(b1, "CT", 'Z', str->l+1, (uint8_t*)str->s);
}
int bam_cigar2ulen(int n_cigar, const uint32_t *cigar)
{
int k, l;
for (k = l = 0; k < n_cigar; ++k) {
if (bam_cigar_type(bam_cigar_op(cigar[k])) &1) {
l += bam_cigar_oplen(cigar[k]);
} else if (bam_cigar_op(cigar[k]) == BAM_CHARD_CLIP) {
l += bam_cigar_oplen(cigar[k]);
}
}
return l;
}
int bam_cigar2matches(int n_cigar, const uint32_t *cigar)
{
int k, l;
for (k = l = 0; k < n_cigar; ++k)
if (bam_cigar_type(bam_cigar_op(cigar[k]))==3)
l += bam_cigar_oplen(cigar[k]);
return l;
}
CigarParser::CigarParser(uint32_t const* cigar, int len)
: cigar_(cigar)
, len_(len)
, readPos_(0)
, refPos_(0)
, currentOpIdx_(0)
, currentOp_(bam_cigar_op(*cigar))
, currentOpLen_(bam_cigar_oplen(*cigar))
, started_(false)
{
}
void calcAlnOffsets(uint32_t *cigar,
uint32_t n_cigar,
uint32_t sa_pos,
char sa_strand,
struct line *l)
{
l->raLen = 0;
l->qaLen = 0;
l->sclip = 0;
l->eclip = 0;
l->SQO = 0;
l->EQO = 0;
bool first = true;
uint32_t k;
for (k = 0; k < n_cigar; ++k)
{
uint32_t opLen = bam_cigar_oplen(cigar[k]);
char opCode = bam_cigar_opchr(cigar[k]);
if (opCode == 'M' || opCode == '=' || opCode == 'X')
{
l->raLen += opLen;
l->qaLen += opLen;
first = false;
}
else if (opCode == 'S' || opCode == 'H')
{
if (first) l->sclip += opLen;
else l->eclip += opLen;
}
else if (opCode == 'D' || opCode == 'N')
{
l->raLen += opLen;
}
else if (opCode == 'I')
{
l->qaLen += opLen;
}
}
//*rapos = str2pos(line->fields[POS]);
l->rapos = sa_pos;
if (sa_strand == '+')
{
l->pos = l->rapos - l->sclip;
l->SQO = l->sclip;
l->EQO = l->sclip + l->qaLen - 1;
}
else
{
l->pos = l->rapos + l->raLen + l->eclip - 1;
l->SQO = l->eclip;
l->EQO = l->eclip + l->qaLen - 1;
}
}
// Adapted from samtools/bam.c
int32_t b2g_bam_clippedlength(bam1_t *b) {
const bam1_core_t c = b->core;
const uint32_t *cigar = bam1_cigar(b);
uint32_t k;
int32_t l = 0;
for (k = 0; k < c.n_cigar; ++k) {
if ('S' == bam_cigar_opchr(bam_cigar_op(cigar[k]))) {
l += bam_cigar_oplen(cigar[k]);
}
}
return l;
}
// Return sum of bases on right of alignment with:
// * hard masked (H)
// * soft masked (S)
// * inserted bases relative to ref (I)
static inline uint32_t bam_get_end_padding(int n_cigar, const uint32_t *cigar)
{
ctx_assert(n_cigar > 0);
uint32_t i, l = 0;
const uint32_t c = (1<<BAM_CINS)|(1<<BAM_CSOFT_CLIP)|(1<<BAM_CHARD_CLIP);
for(i = n_cigar-1; i > 0; i--)
if((c >> bam_cigar_op(cigar[i])) & 1)
l += bam_cigar_oplen(cigar[i]);
return l;
}
/* Make a node containing an InDel
*
* b The input read
* cigar_op_num The operation number of the Insertion/Deletion
*
* returns a node, that must be either inserted into the linked list or
* destroyed with destroyNode()
*/
InDel *makeNode(bam1_t *b, int cigar_op_num) {
int i, op, oplen, quit = 0, type;
int32_t start = b->core.pos-1;
int32_t end;
uint32_t *cigar = bam_get_cigar(b);
InDel *node;
for(i=0; i<cigar_op_num; i++) {
oplen = bam_cigar_oplen(cigar[i]);
type = bam_cigar_type(bam_cigar_op(cigar[i]));
if(type & 2) start += oplen;
}
end = ++start;
for(i=cigar_op_num; i<b->core.n_cigar; i++) {
op = bam_cigar_op(cigar[i]);
oplen = bam_cigar_oplen(cigar[i]);
switch(op) {
case 1: //I
case 2: //D
end = (end>start+oplen) ? end : start+oplen;
break;
default :
quit = 1;
break;
}
if(quit) break;
}
//Make the node
node = calloc(1, sizeof(InDel));
node->tid = b->core.tid;
node->start = start;
node->end = end;
node->count = 1;
return node;
}
/**
* Gets the cigar from a BAM record
*/
void bam_get_cigar_string(bam1_t *s, kstring_t *cigar_string)
{
cigar_string->l=0;
int32_t n_cigar_op = bam_get_n_cigar_op(s);
if (n_cigar_op)
{
uint32_t *cigar = bam_get_cigar(s);
for (int32_t i = 0; i < n_cigar_op; ++i)
{
kputw(bam_cigar_oplen(cigar[i]), cigar_string);
kputc(bam_cigar_opchr(cigar[i]), cigar_string);
}
}
}
void CigarParser::advance() {
int type = bam_cigar_type(currentOp_);
if (type & BAM_CONSUME_REFERENCE) {
refPos_ += currentOpLen_;
}
if (type & BAM_CONSUME_QUERY) {
readPos_ += currentOpLen_;
}
++currentOpIdx_;
assert(currentOpIdx_ < len_);
currentOp_ = bam_cigar_op(cigar_[currentOpIdx_]);
currentOpLen_ = bam_cigar_oplen(cigar_[currentOpIdx_]);
}
unsigned seq_lens_from_bin_cigar (uint32_t* cigar_bin, unsigned cigar_bin_sz, unsigned* q_len, unsigned* r_len)
{
unsigned oplen, constype;
uint32_t *sent;
*q_len = *r_len = 0;
unsigned allen = 0;
for (sent = cigar_bin + cigar_bin_sz; cigar_bin != sent; ++cigar_bin)
{
oplen = bam_cigar_oplen (*cigar_bin);
constype = bam_cigar_type (*cigar_bin);
if (constype & CONSUME_QRY) *q_len += oplen;
if (constype & CONSUME_REF) *r_len += oplen;
allen += oplen;
}
return allen;
}
// Returns 0 on success, -1 on failure.
static int bam_format_cigar(const bam1_t* b, kstring_t* str)
{
// An empty cigar is a special case return "*" rather than ""
if (b->core.n_cigar == 0) {
return (kputc('*', str) == EOF) ? -1 : 0;
}
const uint32_t *cigar = bam_get_cigar(b);
uint32_t i;
for (i = 0; i < b->core.n_cigar; ++i) {
if (kputw(bam_cigar_oplen(cigar[i]), str) == EOF) return -1;
if (kputc(bam_cigar_opchr(cigar[i]), str) == EOF) return -1;
}
return 0;
}
static int32_t unclipped_start(bam1_t *b) {
uint32_t *cigar = bam_get_cigar(b);
int32_t clipped = 0;
uint32_t i;
for (i = 0; i < b->core.n_cigar; i++) {
char c = bam_cigar_opchr(cigar[i]);
if (c == 'S' || c == 'H') { // clips
clipped += bam_cigar_oplen(cigar[i]);
} else {
break;
}
}
return b->core.pos - clipped + 1;
}
// Returns 0 to indicate read should be output 1 otherwise
static int process_aln(const bam_hdr_t *h, bam1_t *b, samview_settings_t* settings)
{
if (settings->remove_B) bam_remove_B(b);
if (settings->min_qlen > 0) {
int k, qlen = 0;
uint32_t *cigar = bam_get_cigar(b);
for (k = 0; k < b->core.n_cigar; ++k)
if ((bam_cigar_type(bam_cigar_op(cigar[k]))&1) || bam_cigar_op(cigar[k]) == BAM_CHARD_CLIP)
qlen += bam_cigar_oplen(cigar[k]);
if (qlen < settings->min_qlen) return 1;
}
if (b->core.qual < settings->min_mapQ || ((b->core.flag & settings->flag_on) != settings->flag_on) || (b->core.flag & settings->flag_off))
return 1;
if (settings->bed && (b->core.tid < 0 || !bed_overlap(settings->bed, h->target_name[b->core.tid], b->core.pos, bam_endpos(b))))
return 1;
if (settings->subsam_frac > 0.) {
uint32_t k = __ac_Wang_hash(__ac_X31_hash_string(bam_get_qname(b)) ^ settings->subsam_seed);
if ((double)(k&0xffffff) / 0x1000000 >= settings->subsam_frac) return 1;
}
if (settings->rghash) {
uint8_t *s = bam_aux_get(b, "RG");
if (s) {
khint_t k = kh_get(rg, settings->rghash, (char*)(s + 1));
if (k == kh_end(settings->rghash)) return 1;
}
}
if (settings->library) {
const char *p = bam_get_library((bam_hdr_t*)h, b);
if (!p || strcmp(p, settings->library) != 0) return 1;
}
if (settings->remove_aux_len) {
size_t i;
for (i = 0; i < settings->remove_aux_len; ++i) {
uint8_t *s = bam_aux_get(b, settings->remove_aux[i]);
if (s) {
bam_aux_del(b, s);
}
}
}
return 0;
}
static void unpad_seq(bam1_t *b, kstring_t *s)
{
int k, j, i;
uint32_t *cigar = bam1_cigar(b);
uint8_t *seq = bam1_seq(b);
ks_resize(s, b->core.l_qseq);
for (k = 0, s->l = 0, j = 0; k < b->core.n_cigar; ++k) {
int op, ol;
op = bam_cigar_op(cigar[k]);
ol = bam_cigar_oplen(cigar[k]);
assert(op == BAM_CMATCH || op == BAM_CDEL || op == BAM_CSOFT_CLIP);
if (op == BAM_CMATCH) {
for (i = 0; i < ol; ++i) s->s[s->l++] = bam1_seqi(seq, j);
++j;
} else if (op == BAM_CSOFT_CLIP) {
j += ol;
} else {
for (i = 0; i < ol; ++i) s->s[s->l++] = 0;
}
}
}
static int unpad_seq(bam1_t *b, kstring_t *s)
{
// Returns 0 on success, -1 on an error
int k, j, i;
int length;
int cigar_n_warning = 0; /* Make this a global and limit to one CIGAR N warning? */
uint32_t *cigar = bam_get_cigar(b);
uint8_t *seq = bam_get_seq(b);
// b->core.l_qseq gives length of the SEQ entry (including soft clips, S)
// We need the padded length after alignment from the CIGAR (excluding
// soft clips S, but including pads from CIGAR D operations)
length = bam_cigar2rlen(b->core.n_cigar, cigar);
ks_resize(s, length);
for (k = 0, s->l = 0, j = 0; k < b->core.n_cigar; ++k) {
int op, ol;
op = bam_cigar_op(cigar[k]);
ol = bam_cigar_oplen(cigar[k]);
if (op == BAM_CMATCH || op == BAM_CEQUAL || op == BAM_CDIFF) {
for (i = 0; i < ol; ++i, ++j) s->s[s->l++] = bam_seqi(seq, j);
} else if (op == BAM_CSOFT_CLIP) {
j += ol;
} else if (op == BAM_CHARD_CLIP) {
/* do nothing */
} else if (op == BAM_CDEL) {
for (i = 0; i < ol; ++i) s->s[s->l++] = 0;
} else if (op == BAM_CREF_SKIP) {
/* Treat CIGAR N as D (not ideal, but better than ignoring it) */
for (i = 0; i < ol; ++i) s->s[s->l++] = 0;
if (0 == cigar_n_warning) {
cigar_n_warning = -1;
fprintf(stderr, "[depad] WARNING: CIGAR op N treated as op D in read %s\n", bam_get_qname(b));
}
} else {
fprintf(stderr, "[depad] ERROR: Didn't expect CIGAR op %c in read %s\n", BAM_CIGAR_STR[op], bam_get_qname(b));
return -1;
}
}
return length != s->l;
}
unsigned alignment_bounds_from_bin_cigar (uint32_t* cigar_bin, unsigned cigar_bin_sz, uint8_t forward, unsigned qry_len, unsigned* q_beg, unsigned* q_end, unsigned* r_beg, unsigned* r_end)
{
unsigned oplen, op, constype;
uint32_t *sent;
*q_beg = *q_end = *r_beg = *r_end = 0;
unsigned allen = 0;
uint32_t tail = 0;
for (sent = cigar_bin + cigar_bin_sz; cigar_bin != sent; ++cigar_bin)
{
oplen = bam_cigar_oplen (*cigar_bin);
op = bam_cigar_op (*cigar_bin);
constype = bam_cigar_type (*cigar_bin);
if (tail && (op == BAM_CHARD_CLIP || op == BAM_CSOFT_CLIP)) // the aligned zone ended, clip started. Note that tail indels are not valid, so we do not assume they are possible..
break;
if (op != BAM_CHARD_CLIP && op != BAM_CSOFT_CLIP)
tail = 1;
if (constype & CONSUME_QRY)
{
if (!tail) *q_beg += oplen;
*q_end += oplen;
}
if (constype & CONSUME_REF)
{
if (!tail) *r_beg += oplen;
*r_end += oplen;
}
allen += oplen;
}
if (!forward)
{
unsigned tmp = qry_len - *q_beg;
*q_beg = qry_len - *q_end;
*q_end = tmp;
}
return allen;
}
static int32_t unclipped_end(bam1_t *b) {
uint32_t *cigar = bam_get_cigar(b);
int32_t end_pos, clipped = 0;
int32_t i;
end_pos = bam_endpos(b);
// now get the clipped end bases (if any)
// if we get to the beginning of the cigar string
// without hitting a non-clip then the results are meaningless
for (i = b->core.n_cigar - 1; i >= 0; i--) {
char c = bam_cigar_opchr(cigar[i]);
if (c == 'S' || c == 'H') { // clips
clipped += bam_cigar_oplen(cigar[i]);
} else {
break;
}
}
return end_pos + clipped;
}
void add_bam_record_to_wiggle(const bam1_t *b, Wiggle& wiggle) {
double w;
if (no_fractional_weight) w = 1.0;
else {
uint8_t *p_tag = bam_aux_get(b, "ZW");
if (p_tag == NULL) return;
w = bam_aux2f(p_tag);
}
int pos = b->core.pos;
uint32_t *p = bam_get_cigar(b);
for (int i = 0; i < (int)b->core.n_cigar; ++i, ++p) {
char op = bam_cigar_op(*p);
int op_len = bam_cigar_oplen(*p);
if (op == BAM_CMATCH)
for (int j = 0; j < op_len; ++j, ++pos) wiggle.read_depth[pos] += w;
else pos += ((bam_cigar_type(op) & 2) ? op_len : 0);
}
}
void cigar_print (FILE* f, uint32_t* cigar, unsigned cigar_sz)
{
uint32_t* sent;
for (sent = cigar+cigar_sz; cigar != sent; ++cigar)
{
uint32_t curop = bam_cigar_op (*cigar);
uint32_t count = bam_cigar_oplen (*cigar);
char schar;
switch (curop)
{
case BAM_CHARD_CLIP:
schar = 'H';
break;
case BAM_CSOFT_CLIP: // skip
schar = 'S';
break;
case BAM_CMATCH:
schar = 'M';
break;
case BAM_CEQUAL:
schar = '=';
break;
case BAM_CDIFF:
schar = '#';
break;
case BAM_CINS:
schar = 'I';
break;
case BAM_CDEL:
schar = 'I';
break;
default:
schar = '?';
}
fprintf (f, "%d%c", count, schar);
}
}
string bam_cigarString (bam1_t *b) {//output CIGAR string
// kstring_t strK;
// kstring_t *str=&strK;
const bam1_core_t *c = &b->core;
string cigarString("");
if ( c->n_cigar > 0 ) {
uint32_t *cigar = bam_get_cigar(b);
for (int i = 0; i < c->n_cigar; ++i) {
cigarString+=to_string((uint)bam_cigar_oplen(cigar[i]))+bam_cigar_opchr(cigar[i]);
};
};
// if (c->n_cigar) { // cigar
// for (int i = 0; i < c->n_cigar; ++i) {
// kputw(bam_cigar_oplen(cigar[i]), str);
// kputc(bam_cigar_opchr(cigar[i]), str);
// }
// } else kputc('*', str);
//
// string cigarString (str->s,str->l);
return cigarString;
};
// prepare internal structures for clipping and alignment
// returns true if realignment was performed
bool RealignImp::compute_alignment (
const char* q_seq,
unsigned q_len,
const char* r_seq,
unsigned r_len,
int r_pos,
bool forward,
const uint32_t* cigar,
unsigned cigar_sz,
uint32_t*& cigar_dest,
unsigned& cigar_dest_sz,
int& new_pos,
bool& already_perfect,
bool& clip_failed,
bool& alignment_failed,
bool& unclip_failed)
{
already_perfect = false;
alignment_failed = false;
unclip_failed = false;
unsigned oplen;
const char* q_seq_clipped = q_seq;
const uint32_t* cigar_clipped = cigar;
unsigned cigar_sz_clipped = cigar_sz;
unsigned sclip_q_len, sclip_r_len, sclip_al_len;
assert (cigar_sz);
// reset realigner
Reset ();
// set clipping
SetClipping ((int) cliptype_, forward);
// clip out the hard and soft clipping zones from 5" and 3"
// The 'cut out' of the q_seq is done by switching to downstream pointer.
if (bam_cigar_op (*cigar) == BAM_CSOFT_CLIP)
{
oplen = bam_cigar_oplen (*cigar);
ClipStart (oplen);
q_seq_clipped += oplen;
++cigar_clipped;
--cigar_sz_clipped;
}
if (cigar_sz > 1 && bam_cigar_op (cigar [cigar_sz - 1]) == BAM_CSOFT_CLIP)
{
oplen = bam_cigar_oplen (cigar [cigar_sz - 1]);
ClipEnd (oplen);
--cigar_sz_clipped;
}
// cigar defines q_seq and t_seq lengths
sclip_al_len = seq_lens_from_bin_cigar (cigar_clipped, cigar_sz_clipped, &sclip_q_len, &sclip_r_len);
const std::string query (q_seq_clipped, sclip_q_len);
const std::string target (r_seq, sclip_r_len);
std::string pretty_al; pretty_al.reserve (sclip_al_len);
pretty_al_from_bin_cigar (cigar_clipped, cigar_sz_clipped, q_seq_clipped, r_seq, pretty_al);
// Realigner requires strings of proper size to be passed to SetSequences
SetSequences (query, target, pretty_al, forward);
if (!ClipAnchors (clip_failed))
{
already_perfect = true;
return false; // alignment already good, no imperfect zone to realign found
}
// TODO avoid automatic vectors to prevent unneeded heap usage
vector<MDelement> new_md_vec;
vector<CigarOp> new_cigar_vec;
unsigned int start_pos_shift;
if (!computeSWalignment(new_cigar_vec, new_md_vec, start_pos_shift))
{
alignment_failed = true;
return false;
}
if (!addClippedBasesToTags(new_cigar_vec, new_md_vec, q_len))
{
unclip_failed = true;
return false; // error adding back clipped out zones
}
if (!LeftAnchorClipped () && start_pos_shift != 0)
{
// build cigar data only if it is needed
// TODO avoid automatic vectors to prevent unneeded heap usage
std::vector <CigarOp> cigar_vec;
cigar_vector_from_bin (cigar, cigar_sz, cigar_vec);
new_pos = updateReadPosition (cigar_vec, start_pos_shift, r_pos);
}
else
new_pos = r_pos;
// free (cigar_dest);
// TODO: switch to better alignment memory management, avoid heap operations
cigar_dest = (uint32_t*) tmap_malloc (sizeof (uint32_t) * new_cigar_vec.size (), "cigar_dest");
cigar_dest_sz = new_cigar_vec.size ();
cigar_vector_to_bin (new_cigar_vec, cigar_dest);
return true;
}
static aln_t align_read_against_one(kseq_t *target, const int read_len,
uint8_t *read_num, kswq_t **qry,
const align_config_t *conf,
const int min_score) {
uint8_t *ref_num = calloc(target->seq.l, sizeof(uint8_t));
for (size_t k = 0; k < target->seq.l; ++k)
ref_num[k] = conf->table[(int)target->seq.s[k]];
aln_t aln;
aln.cigar = NULL;
aln.loc = ksw_align(read_len, read_num, target->seq.l, ref_num, conf->m,
conf->mat, conf->gap_o, conf->gap_e, KSW_XSTART, qry);
aln.target_name = target->name.s;
if (aln.loc.score < min_score) {
free(ref_num);
return aln;
}
ksw_global(aln.loc.qe - aln.loc.qb + 1, &read_num[aln.loc.qb],
aln.loc.te - aln.loc.tb + 1, &ref_num[aln.loc.tb], conf->m,
conf->mat, conf->gap_o, conf->gap_e, conf->bandwidth, &aln.n_cigar,
&aln.cigar);
aln.nm = 0;
size_t qi = aln.loc.qb, ri = aln.loc.tb;
for (int k = 0; k < aln.n_cigar; k++) {
const int32_t oplen = bam_cigar_oplen(aln.cigar[k]),
optype = bam_cigar_type(aln.cigar[k]);
if (optype & 3) { // consumes both - check for mismatches
for (int j = 0; j < oplen; j++) {
if (UNLIKELY(read_num[qi + j] != ref_num[ri + j]))
aln.nm++;
}
} else {
aln.nm += oplen;
}
if (optype & 1)
qi += oplen;
if (optype & 2)
ri += oplen;
}
free(ref_num);
/* size_t cigar_len = aln.loc.qb; */
/* for (int c = 0; c < aln.n_cigar; c++) { */
/* int32_t length = (0xfffffff0 & *(aln.cigar + c)) >> 4; */
/* cigar_len += length; */
/* } */
/* cigar_len += read_len - aln.loc.qe - 1; */
/* if(cigar_len != (size_t)read_len) { */
/* /\* printf("[ig_align] Error: cigar length (score %d) not equal to read length for XXX (target %s): %zu vs %d\n", aln.loc.score, target->name.s, cigar_len, read_len); *\/ */
/* // NOTE: */
/* // It is *really* *f*****g* *scary* that it's spitting out cigars that are not the same length as the query sequence. */
/* // Nonetheless, fixing it seems to involve delving into the depths of ksw_align() and ksw_global(), which would be very time consuming, and the length discrepancy seems to ony appear in very poor matches. */
/* // I.e., poor enough that we will subsequently ignore them in partis/python/waterer.py, so it seems to not screw anything up downstream to just set the length-discrepant matches' scores to zero, such that ig-sw doesn't write them to its sam output. */
/* // Note also that it is not always the lowest- or highest-scoring matches that have discrepant lengths (i.e. setting their scores to zero promotes matches swith poorer scores, but which do not have discrepant lengths. */
/* /\* aln.loc.score = 0; *\/ */
/* aln.cigar = NULL; */
/* } */
return aln;
}
static aln_v align_read(const kseq_t *read,
const kseq_v targets,
const align_config_t *conf)
{
kseq_t *r;
const int32_t read_len = read->seq.l;
aln_v result;
kv_init(result);
kv_resize(aln_t, result, kv_size(targets));
uint8_t *read_num = calloc(read_len, sizeof(uint8_t));
for(size_t k = 0; k < read_len; ++k)
read_num[k] = conf->table[(int)read->seq.s[k]];
// Align to each target
kswq_t *qry = NULL;
for(size_t j = 0; j < kv_size(targets); j++) {
// Encode target
r = &kv_A(targets, j);
uint8_t *ref_num = calloc(r->seq.l, sizeof(uint8_t));
for(size_t k = 0; k < r->seq.l; ++k)
ref_num[k] = conf->table[(int)r->seq.s[k]];
aln_t aln;
aln.target_idx = j;
aln.loc = ksw_align(read_len, read_num,
r->seq.l, ref_num,
conf->m,
conf->mat,
conf->gap_o,
conf->gap_e,
KSW_XSTART,
&qry);
ksw_global(aln.loc.qe - aln.loc.qb + 1,
&read_num[aln.loc.qb],
aln.loc.te - aln.loc.tb + 1,
&ref_num[aln.loc.tb],
conf->m,
conf->mat,
conf->gap_o,
conf->gap_e,
50, /* TODO: Magic number - band width */
&aln.n_cigar,
&aln.cigar);
aln.nm = 0;
size_t qi = aln.loc.qb, ri = aln.loc.tb;
for(size_t k = 0; k < aln.n_cigar; k++) {
const int32_t oplen = bam_cigar_oplen(aln.cigar[k]),
optype = bam_cigar_type(aln.cigar[k]);
if(optype & 3) { // consumes both - check for mismatches
for(size_t j = 0; j < oplen; j++) {
if(UNLIKELY(read_num[qi + j] != ref_num[ri + j]))
aln.nm++;
}
} else {
aln.nm += oplen;
}
if(optype & 1) qi += oplen;
if(optype & 2) ri += oplen;
}
kv_push(aln_t, result, aln);
free(ref_num);
}
free(qry);
free(read_num);
ks_introsort(dec_score, kv_size(result), result.a);
return result;
}
int bam_pad2unpad(samFile *in, samFile *out, bam_hdr_t *h, faidx_t *fai)
{
bam1_t *b = 0;
kstring_t r, q;
int r_tid = -1;
uint32_t *cigar2 = 0;
int ret = 0, n2 = 0, m2 = 0, *posmap = 0;
b = bam_init1();
r.l = r.m = q.l = q.m = 0; r.s = q.s = 0;
int read_ret;
while ((read_ret = sam_read1(in, h, b)) >= 0) { // read one alignment from `in'
// Cannot depad unmapped CRAM data
if (b->core.flag & BAM_FUNMAP)
goto next_seq;
uint32_t *cigar = bam_get_cigar(b);
n2 = 0;
if (b->core.pos == 0 && b->core.tid >= 0 && strcmp(bam_get_qname(b), h->target_name[b->core.tid]) == 0) {
// fprintf(stderr, "[depad] Found embedded reference '%s'\n", bam_get_qname(b));
r_tid = b->core.tid;
if (0!=unpad_seq(b, &r)) {
fprintf(stderr, "[depad] ERROR: Problem parsing SEQ and/or CIGAR in reference %s\n", bam_get_qname(b));
return -1;
};
if (h->target_len[r_tid] != r.l) {
fprintf(stderr, "[depad] ERROR: (Padded) length of '%s' is %u in BAM header, but %llu in embedded reference\n", bam_get_qname(b), h->target_len[r_tid], (unsigned long long)(r.l));
return -1;
}
if (fai) {
// Check the embedded reference matches the FASTA file
if (load_unpadded_ref(fai, h->target_name[b->core.tid], h->target_len[b->core.tid], &q)) {
fprintf(stderr, "[depad] ERROR: Failed to load embedded reference '%s' from FASTA\n", h->target_name[b->core.tid]);
return -1;
}
assert(r.l == q.l);
int i;
for (i = 0; i < r.l; ++i) {
if (r.s[i] != q.s[i]) {
// Show gaps as ASCII 45
fprintf(stderr, "[depad] ERROR: Embedded sequence and reference FASTA don't match for %s base %i, '%c' vs '%c'\n",
h->target_name[b->core.tid], i+1,
r.s[i] ? seq_nt16_str[(int)r.s[i]] : 45,
q.s[i] ? seq_nt16_str[(int)q.s[i]] : 45);
return -1;
}
}
}
write_cigar(cigar2, n2, m2, bam_cigar_gen(b->core.l_qseq, BAM_CMATCH));
replace_cigar(b, n2, cigar2);
posmap = update_posmap(posmap, r);
} else if (b->core.n_cigar > 0) {
int i, k, op;
if (b->core.tid < 0) {
fprintf(stderr, "[depad] ERROR: Read '%s' has CIGAR but no RNAME\n", bam_get_qname(b));
return -1;
} else if (b->core.tid == r_tid) {
; // good case, reference available
//fprintf(stderr, "[depad] Have ref '%s' for read '%s'\n", h->target_name[b->core.tid], bam_get_qname(b));
} else if (fai) {
if (load_unpadded_ref(fai, h->target_name[b->core.tid], h->target_len[b->core.tid], &r)) {
fprintf(stderr, "[depad] ERROR: Failed to load '%s' from reference FASTA\n", h->target_name[b->core.tid]);
return -1;
}
posmap = update_posmap(posmap, r);
r_tid = b->core.tid;
// fprintf(stderr, "[depad] Loaded %s from FASTA file\n", h->target_name[b->core.tid]);
} else {
fprintf(stderr, "[depad] ERROR: Missing %s embedded reference sequence (and no FASTA file)\n", h->target_name[b->core.tid]);
return -1;
}
if (0!=unpad_seq(b, &q)) {
fprintf(stderr, "[depad] ERROR: Problem parsing SEQ and/or CIGAR in read %s\n", bam_get_qname(b));
return -1;
};
if (bam_cigar_op(cigar[0]) == BAM_CSOFT_CLIP) {
write_cigar(cigar2, n2, m2, cigar[0]);
} else if (bam_cigar_op(cigar[0]) == BAM_CHARD_CLIP) {
write_cigar(cigar2, n2, m2, cigar[0]);
if (b->core.n_cigar > 2 && bam_cigar_op(cigar[1]) == BAM_CSOFT_CLIP) {
write_cigar(cigar2, n2, m2, cigar[1]);
}
}
/* Determine CIGAR operator for each base in the aligned read */
for (i = 0, k = b->core.pos; i < q.l; ++i, ++k)
q.s[i] = q.s[i]? (r.s[k]? BAM_CMATCH : BAM_CINS) : (r.s[k]? BAM_CDEL : BAM_CPAD);
/* Include any pads if starts with an insert */
if (q.s[0] == BAM_CINS) {
for (k = 0; k+1 < b->core.pos && !r.s[b->core.pos - k - 1]; ++k);
if (k) write_cigar(cigar2, n2, m2, bam_cigar_gen(k, BAM_CPAD));
k = 0;
} else if (q.s[0] == BAM_CPAD) {
// Join 'k' CPAD to our first cigar op CPAD too.
for (k = 0; k+1 < b->core.pos && !r.s[b->core.pos - k - 1]; ++k);
} else {
k = 0;
}
/* Count consecutive CIGAR operators to turn into a CIGAR string */
for (i = 1, k++, op = q.s[0]; i < q.l; ++i) {
if (op != q.s[i]) {
write_cigar(cigar2, n2, m2, bam_cigar_gen(k, op));
op = q.s[i]; k = 1;
} else ++k;
}
write_cigar(cigar2, n2, m2, bam_cigar_gen(k, op));
if (bam_cigar_op(cigar[b->core.n_cigar-1]) == BAM_CSOFT_CLIP) {
write_cigar(cigar2, n2, m2, cigar[b->core.n_cigar-1]);
} else if (bam_cigar_op(cigar[b->core.n_cigar-1]) == BAM_CHARD_CLIP) {
if (b->core.n_cigar > 2 && bam_cigar_op(cigar[b->core.n_cigar-2]) == BAM_CSOFT_CLIP) {
write_cigar(cigar2, n2, m2, cigar[b->core.n_cigar-2]);
}
write_cigar(cigar2, n2, m2, cigar[b->core.n_cigar-1]);
}
/* Remove redundant P operators between M/X/=/D operators, e.g. 5M2P10M -> 15M */
int pre_op, post_op;
for (i = 2; i < n2; ++i)
if (bam_cigar_op(cigar2[i-1]) == BAM_CPAD) {
pre_op = bam_cigar_op(cigar2[i-2]);
post_op = bam_cigar_op(cigar2[i]);
/* Note don't need to check for X/= as code above will use M only */
if ((pre_op == BAM_CMATCH || pre_op == BAM_CDEL) && (post_op == BAM_CMATCH || post_op == BAM_CDEL)) {
/* This is a redundant P operator */
cigar2[i-1] = 0; // i.e. 0M
/* If had same operator either side, combine them in post_op */
if (pre_op == post_op) {
/* If CIGAR M, could treat as simple integers since BAM_CMATCH is zero*/
cigar2[i] = bam_cigar_gen(bam_cigar_oplen(cigar2[i-2]) + bam_cigar_oplen(cigar2[i]), post_op);
cigar2[i-2] = 0; // i.e. 0M
}
}
}
/* Remove the zero'd operators (0M) */
for (i = k = 0; i < n2; ++i)
if (cigar2[i]) cigar2[k++] = cigar2[i];
n2 = k;
replace_cigar(b, n2, cigar2);
}
/* Even unmapped reads can have a POS value, e.g. if their mate was mapped */
if (b->core.pos != -1) b->core.pos = posmap[b->core.pos];
if (b->core.mtid < 0 || b->core.mpos < 0) {
/* Nice case, no mate to worry about*/
// fprintf(stderr, "[depad] Read '%s' mate not mapped\n", bam_get_qname(b));
/* TODO - Warning if FLAG says mate should be mapped? */
/* Clean up funny input where mate position is given but mate reference is missing: */
b->core.mtid = -1;
b->core.mpos = -1;
} else if (b->core.mtid == b->core.tid) {
/* Nice case, same reference */
// fprintf(stderr, "[depad] Read '%s' mate mapped to same ref\n", bam_get_qname(b));
b->core.mpos = posmap[b->core.mpos];
} else {
/* Nasty case, Must load alternative posmap */
// fprintf(stderr, "[depad] Loading reference '%s' temporarily\n", h->target_name[b->core.mtid]);
if (!fai) {
fprintf(stderr, "[depad] ERROR: Needed reference %s sequence for mate (and no FASTA file)\n", h->target_name[b->core.mtid]);
return -1;
}
/* Temporarily load the other reference sequence */
if (load_unpadded_ref(fai, h->target_name[b->core.mtid], h->target_len[b->core.mtid], &r)) {
fprintf(stderr, "[depad] ERROR: Failed to load '%s' from reference FASTA\n", h->target_name[b->core.mtid]);
return -1;
}
posmap = update_posmap(posmap, r);
b->core.mpos = posmap[b->core.mpos];
/* Restore the reference and posmap*/
if (load_unpadded_ref(fai, h->target_name[b->core.tid], h->target_len[b->core.tid], &r)) {
fprintf(stderr, "[depad] ERROR: Failed to load '%s' from reference FASTA\n", h->target_name[b->core.tid]);
return -1;
}
posmap = update_posmap(posmap, r);
}
/* Most reads will have been moved so safest to always recalculate the BIN value */
b->core.bin = bam_reg2bin(b->core.pos, bam_endpos(b));
next_seq:
sam_write1(out, h, b);
}
if (read_ret < -1) {
fprintf(stderr, "[depad] truncated file.\n");
ret = 1;
}
free(r.s); free(q.s); free(posmap);
bam_destroy1(b);
return ret;
}
int bsstrand_func(bam1_t *b, const samfile_t *in, samfile_t *out, void *data) {
bsstrand_data_t *d = (bsstrand_data_t*)data;
bsstrand_conf_t *conf = d->conf;
const bam1_core_t *c = &b->core;
if (c->flag & BAM_FUNMAP){
if (out) samwrite(out, b);
d->n_unmapped++;
return 0;
}
fetch_refseq(d->rs, in->header->target_name[c->tid], c->pos, c->pos+1);
uint32_t rpos=c->pos+1, qpos=0;
int i, nC2T = 0, nG2A = 0;
uint32_t j;
char rbase, qbase;
for (i=0; i<c->n_cigar; ++i) {
uint32_t op = bam_cigar_op(bam1_cigar(b)[i]);
uint32_t oplen = bam_cigar_oplen(bam1_cigar(b)[i]);
switch(op) {
case BAM_CMATCH:
for(j=0; j<oplen; ++j) {
rbase = toupper(getbase_refseq(d->rs, rpos+j));
qbase = bscall(bam1_seq(b), qpos+j);
if (rbase == 'C' && qbase == 'T') nC2T += 1;
if (rbase == 'G' && qbase == 'A') nG2A += 1;
/* printf("%c vs %c\n", toupper(rbase), qbase); */
}
rpos += oplen;
qpos += oplen;
break;
case BAM_CINS:
qpos += oplen;
break;
case BAM_CDEL:
rpos += oplen;
break;
case BAM_CSOFT_CLIP:
qpos += oplen;
break;
default:
fprintf(stderr, "Unknown cigar, %u\n", op);
abort();
}
}
char key[2] = {'Z','S'};
unsigned char *bsstrand = bam_aux_get(b, key);
if (bsstrand) {
bsstrand++;
double s = similarity(nG2A, nC2T);
if (nG2A > 1 && nC2T > 1 && s > 0.5) {
if (conf->output_read || conf->output_all_read)
printf("F\t%s\t%d\t%d\t%d\t%s\t%s\t%1.2f\n", in->header->target_name[c->tid], c->pos, nC2T, nG2A, bam1_qname(b), bsstrand, s);
bam_aux_append(b, "OS", 'A', 1, bsstrand);
bsstrand[0] = '?';
d->n_fail++;
} else if (*bsstrand == '+' && nG2A > nC2T + 2) {
if (conf->output_read || conf->output_all_read)
printf("W2C\t%s\t%d\t%d\t%d\t%s\t%s\t%1.2f\n", in->header->target_name[c->tid], c->pos, nC2T, nG2A, bam1_qname(b), bsstrand, s);
bam_aux_append(b, "OS", 'A', 1, bsstrand);
bsstrand[0] = '-';
d->n_corr++;
} else if (*bsstrand == '-' && nC2T > nG2A + 2) {
if (conf->output_read || conf->output_all_read)
printf("C2W\t%s\t%d\t%d\t%d\t%s\t%s\t%1.2f\n", in->header->target_name[c->tid], c->pos, nC2T, nG2A, bam1_qname(b), bsstrand, s);
bam_aux_append(b, "OS", 'A', 1, bsstrand);
bsstrand[0] = '+';
d->n_corr++;
} else if (conf->output_all_read) {
printf("N\t%s\t%d\t%d\t%d\t%s\t%s\t%1.2f\n", in->header->target_name[c->tid], c->pos, nC2T, nG2A, bam1_qname(b), bsstrand, s);
}
} else if (!(c->flag & BAM_FUNMAP) && conf->infer_bsstrand) {
char bss[3];
if (similarity(nG2A, nC2T) < 0.5) {
strcpy(bss, "??");
} else if (nC2T > nG2A) {
strcpy(bss, c->flag & BAM_FREVERSE ? "+-" : "++");
} else {
strcpy(bss, c->flag & BAM_FREVERSE ? "-+" : "--");
}
bam_aux_append(b, "ZS", 'Z', 3, (uint8_t*) bss);
}
if (out) samwrite(out, b);
d->n_mapped++;
return 0;
}
/**
* Gets the base in the read that is mapped to a genomic position.
* Extracts the read sequence and aualities too.
*/
void bam_get_base_and_qual_and_read_and_qual(bam1_t *srec, uint32_t pos, char& base, char& qual, int32_t& rpos, kstring_t* readseq, kstring_t* readqual)
{
bam1_core_t *c = &srec->core;
int32_t rlen = c->l_qseq;
uint32_t cpos = c->pos; //reference coordinates of the first mapped base
rpos = 0; //read coordinates
kstring_t str;
str.l = str.m = 0, str.s = 0;
base = 'N';
qual = 0;
if (c->n_cigar)
{
uint32_t *cigar = bam_get_cigar(srec);
for (uint32_t i = 0; i < c->n_cigar; ++i)
{
char op = bam_cigar_opchr(cigar[i]);
str.l = 0;
kputw(bam_cigar_oplen(cigar[i]), &str);
char* stop;
uint32_t len = strtol(str.s, &stop, 10);
assert(stop);
if (op=='M')
{
if (pos>=cpos && pos<=cpos+len-1)
{
rpos += pos-cpos;
break;
}
cpos += len;
rpos += len;
}
else if (op=='D')
{
if (pos>=cpos && pos<=cpos+len-1)
{
rpos = -1;
break;
}
cpos += len;
}
else if (op=='S' || op=='I')
{
rpos += len;
}
}
//std::cout << "bpos " << bpos << "\n";
if (rpos>=0 && rpos<=rlen)
{
//sequence
bam_get_seq_string(srec, readseq);
base = readseq->s[rpos];
//qual
bam_get_qual_string(srec, readqual);
qual = readqual->s[rpos];
}
else
{
rpos = BAM_READ_INDEX_NA;
}
}
// std::cout << "b: " << base << "\n";
// std::cout << "q: " << s[bpos-1] << " " << q << "\n";
// for (uint32_t i = 0; i < c->l_qseq; ++i) std::cerr << ((char)(s[i] + 33));
};
static int trim_ns(bam1_t *b, void *data) {
int ret = 0;
opts_t *op((opts_t *)data);
std::vector<uint8_t> aux(bam_get_aux(b), bam_get_aux(b) + bam_get_l_aux(b));
int tmp;
uint8_t *const seq(bam_get_seq(b));
uint32_t *const cigar(bam_get_cigar(b));
//op->n_cigar = b->core.n_cigar;
op->resize(b->l_data); // Make sure it's big enough to hold everything.
memcpy(op->data, b->data, b->core.l_qname);
// Get #Ns at the beginning
for(tmp = 0; bam_seqi(seq, tmp) == dlib::htseq::HTS_N; ++tmp);
const int n_start(tmp);
if(tmp == b->core.l_qseq - 1) // all bases are N -- garbage read
ret |= op->skip_all_ns;
// Get #Ns at the end
for(tmp = b->core.l_qseq - 1; bam_seqi(seq, tmp) == dlib::htseq::HTS_N; --tmp);
const int n_end(b->core.l_qseq - 1 - tmp);
// Get new length for read
int final_len(b->core.l_qseq - n_end - n_start);
if(final_len < 0) final_len = 0;
if(final_len < op->min_trimmed_len) // Too short.
ret |= 1;
// Copy in qual and all of aux.
if(n_end) {
if((tmp = bam_cigar_oplen(cigar[b->core.n_cigar - 1]) - n_end) == 0) {
LOG_DEBUG("Entire cigar operation is the softclip. Decrease the number of new cigar operations.\n");
--b->core.n_cigar;
} else {
LOG_DEBUG("Updating second cigar operation in-place.\n");
cigar[b->core.n_cigar - 1] = bam_cigar_gen(tmp, BAM_CSOFT_CLIP);
}
}
// Get new n_cigar.
if((tmp = bam_cigar_oplen(*cigar) - n_start) == 0) {
memcpy(op->data + b->core.l_qname, cigar + 1, (--b->core.n_cigar) << 2); // << 2 for 4 bit per cigar op
} else {
if(n_start) *cigar = bam_cigar_gen(tmp, BAM_CSOFT_CLIP);
memcpy(op->data + b->core.l_qname, cigar, b->core.n_cigar << 2);
}
uint8_t *opseq(op->data + b->core.l_qname + (b->core.n_cigar << 2)); // Pointer to the seq region of new data field.
for(tmp = 0; tmp < final_len >> 1; ++tmp)
opseq[tmp] = (bam_seqi(seq, ((tmp << 1) + n_start)) << 4) | (bam_seqi(seq, (tmp << 1) + n_start + 1));
if(final_len & 1)
opseq[tmp] = (bam_seqi(seq, ((tmp << 1) + n_start)) << 4);
tmp = bam_get_l_aux(b);
memcpy(opseq + ((final_len + 1) >> 1), bam_get_qual(b) + n_start, final_len + tmp);
// Switch data strings
std::swap(op->data, b->data);
b->core.l_qseq = final_len;
memcpy(bam_get_aux(b), aux.data(), aux.size());
b->l_data = (bam_get_aux(b) - b->data) + aux.size();
if(n_end) bam_aux_append(b, "NE", 'i', sizeof(int), (uint8_t *)&n_end);
if(n_start) bam_aux_append(b, "NS", 'i', sizeof(int), (uint8_t *)&n_start);
const uint32_t *pvar((uint32_t *)dlib::array_tag(b, "PV"));
tmp = b->core.flag & BAM_FREVERSE ? n_end: n_start;
if(pvar) {
std::vector<uint32_t>pvals(pvar + tmp, pvar + final_len + tmp);
bam_aux_del(b, (uint8_t *)(pvar) - 6);
dlib::bam_aux_array_append(b, "PV", 'I', sizeof(uint32_t), final_len, (uint8_t *)pvals.data());
}
const uint32_t *fvar((uint32_t *)dlib::array_tag(b, "FA"));
if(fvar) {
std::vector<uint32_t>fvals(fvar + tmp, fvar + final_len + tmp);
bam_aux_del(b, (uint8_t *)(fvar) - 6);
dlib::bam_aux_array_append(b, "FA", 'I', sizeof(uint32_t), final_len, (uint8_t *)fvals.data());
}
return ret;
}
int do_grep() {
#ifdef DEBUGa
printf("[!]do_grep\n");
#endif
BamInfo_t *pbam;
kh_cstr_t BamID;
khiter_t ki, bami;
kstring_t ks1 = { 0, 0, NULL };
kstring_t ks2 = { 0, 0, NULL };
kstring_t ks3 = { 0, 0, NULL };
samFile *in;
bam_hdr_t *h;
hts_idx_t *idx;
bam1_t *b, *d, *d2, *bR1, *bR2, *bR3;
bR1 = bam_init1(); bR2 = bam_init1(); bR3 = bam_init1();
//htsFile *out;
//hts_opt *in_opts = NULL, *out_opts = NULL;
int r = 0, exit_code = 0;
kvec_t(bam1_t) R1, R2, RV;
pierCluster_t *pierCluster;
//samdat_t tmp_samdat;
FILE *fs = fopen("./test.txt","w");
for (bami = kh_begin(bamNFOp); bami != kh_end(bamNFOp); ++bami) {
//printf(">[%d]:\n",bami);
if (kh_exist(bamNFOp, bami)) {
kv_init(R1); kv_init(R2); kv_init(RV);
//tmp_samdat = (const samdat_t){ 0 };
//memset(&tmp_samdat,0,sizeof(samdat_t));
//printf("-[%d]:\n",bami);
BamID = kh_key(bamNFOp, bami);
pbam = &kh_value(bamNFOp, bami);
fprintf(stderr, "%u [%s]=%s\t%u %u\n",bami,BamID,pbam->fileName,pbam->insertSize,pbam->SD);
in = sam_open(pbam->fileName, "r");
if (in == NULL) {
fprintf(stderr, "[x]Error opening \"%s\"\n", pbam->fileName);
return EXIT_FAILURE;
}
h = sam_hdr_read(in);
/* out = hts_open("-", "w");
if (out == NULL) {
fprintf(stderr, "[x]Error opening standard output\n");
return EXIT_FAILURE;
}
if (sam_hdr_write(out, h) < 0) {
fprintf(stderr, "[!]Error writing output header.\n");
exit_code = 1;
}
*/
int8_t *ChrIsHum;
if (h == NULL) {
fprintf(stderr, "[x]Couldn't read header for \"%s\"\n", pbam->fileName);
return EXIT_FAILURE;
} else {
ChrIsHum = malloc(h->n_targets * sizeof(int8_t));
for (int32_t i=0; i < h->n_targets; ++i) {
//ChrIsHum[i] = -1;
ki = kh_get(chrNFO, chrNFOp, h->target_name[i]);
if (ki == kh_end(chrNFOp)) {
errx(4,"[x]Cannot find ChrID for [%s] !",h->target_name[i]);
} else {
ChrInfo_t * tmp = &kh_value(chrNFOp, ki);
ChrIsHum[i] = tmp->isHum;
//printf(">>> %d Chr:%s %d\n",i,h->target_name[i],ChrIsHum[i]);
}
}
}
h->ignore_sam_err = 0;
b = bam_init1();
d = bam_init1();
d2 = bam_init1();
if ((idx = sam_index_load(in, pbam->fileName)) == 0) {
fprintf(stderr, "[E::%s] fail to load the BAM index\n", __func__);
return 1;
}
pierCluster = sam_plp_init();
while ((r = sam_read1(in, h, b)) >= 0) {
int8_t flag = false;
const bam1_core_t *c = &b->core;
if (c->flag & BAM_FSECONDARY) continue;
if (c->n_cigar) {
uint32_t *cigar = bam_get_cigar(b);
for (int i = 0; i < c->n_cigar; ++i) {
if (bam_cigar_opchr(cigar[i])=='S') { // soft clipping
if ( bam_cigar_oplen(cigar[i]) >= myConfig.minGrepSlen ) {
flag = true;
}
}
}
}
if (flag && ChrIsHum[c->tid]) { // Now, skip Virus items.
//bam_copy1(bR1, b);
flag = 0; // recycle
//int enoughMapQ = 0;
//kstring_t ks = { 0, 0, NULL };
/*if (sam_format1(h, b, &ks1) < 0) {
fprintf(stderr, "Error writing output.\n");
exit_code = 1;
break;
} else*/ if ((c->mtid == c->tid && ChrIsHum[c->tid]) || (ChrIsHum[c->tid] ^ ChrIsHum[c->mtid])) { // Only grep those mapped on same Human ChrID, or diff species/一方在病毒的情况.
//printf(">[%s]\n",ks_str(&ks1));
flag |= 1;
//tmp_samdat.b = bam_dup1(b);
//kv_push(samdat_t,R1,tmp_samdat);
/*if (checkMapQ(ChrIsHum, b, true)) {
++enoughMapQ;
}*/
}
if (getPairedSam(in, idx, b, d) != 0) {
flag &= ~1;
continue;
} else {
flag |= 2;
/*if (checkMapQ(ChrIsHum, d, false)) {
++enoughMapQ;
}*/
/*if (c->flag & BAM_FSECONDARY) {
if (getPairedSam(in, idx, d, d2) == 0) {
//sam_format1(h, d2, &ks3);
flag |= 4;
if (checkMapQ(ChrIsHum, d2, false)) {
++enoughMapQ;
}
}
}*/
}
/*
对于 BAM_FSECONDARY(256) 的 Read，跳两次 与 读 SA 项，效果一样。
>[sf95_Ref_48245009_48245108_48245208_Vir_-_2000_2044_R_100_90 353 chr2 13996555 0 50S40M chr18 48245109 0ACACAACAATGTTCCGGAGACTCTAAGGCCTCCCGATACAGAGCAGAGGCCACACACACACACACCATGGAATACTATTCAGCCAAAAAA CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCC NM:i:0 MD:Z:40 AS:i:40 XS:i:40 RG:Z:Fsimout_mB SA:Z:rgi|59585|emb|X04615.1|,2000,-,40S46M4S,60,0; YC:Z:CT YD:Z:f]
-[sf95_Ref_48245009_48245108_48245208_Vir_-_2000_2044_R_100_90 177 chr18 48245109 9 40S50M gi|59585|emb|X04615.1|2000 0 GTTCCGGAGACTCTAAGGCCTCCCGATACAGAGCAGAGGCCACACACACACACACCATGGAATACTATTCAGCCAAAAAAAGGAATTCAA CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCC NM:i:0 MD:Z:50 AS:i:50 XS:i:46 RG:Z:Fsimout_mB SA:Z:rgi|59585|emb|X04615.1|,2000,+,50S40M,9,0; YC:Z:GA YD:Z:f]
+[sf95_Ref_48245009_48245108_48245208_Vir_-_2000_2044_R_100_90 113 gi|59585|emb|X04615.1| 2000 60 40S46M4S chr18 48245109 0 TTTTTTGGCTGAATAGTATTCCATGGTGTGTGTGTGTGTGGCCTCTGCTCTGTATCGGGAGGCCTTAGAGTCTCCGGAACATTGTTGTGT CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCC NM:i:0 MD:Z:46 AS:i:46 XS:i:27 RG:Z:Fsimout_mB SA:Z:fchr2,13996555,+,50S40M,0,0; YC:Z:CT YD:Z:r]
*/
/*if (sam_format1(h, d, &ks2) < 0) {
fprintf(stderr, "Error writing output.\n");
exit_code = 1;
break;
}*/
if (((flag & 3) == 3) /*&& enoughMapQ >= myConfig.samples*/) {
/*printf(">%d[%s]\n",checkMapQ(ChrIsHum, b, true),ks_str(&ks1));
printf("-%d[%s]\n",checkMapQ(ChrIsHum, d, false),ks_str(&ks2));
if (flag & 4) {
printf("+%d[%s]\n",checkMapQ(ChrIsHum, d2, false),ks_str(&ks3));
}
printf("<--%d\n",enoughMapQ);*/
if (sam_plp_push(ChrIsHum, pierCluster, b) == 0) {
//printf("--HumRange=%s:%d-%d\n", h->target_name[(pierCluster->HumanRange).tid], (pierCluster->HumanRange).pos, (pierCluster->HumanRange).endpos);
if ((!ChrIsHum[(d->core).tid]) && (flag & 2)) sam_plp_push(ChrIsHum, pierCluster, d);
//if ((!ChrIsHum[(d2->core).tid]) && (flag & 4)) sam_plp_push(ChrIsHum, pierCluster, d2);
} else {
//print
fprintf(fs,"[%s]\nHumRange=%s:%d-%d\n", BamID, h->target_name[(pierCluster->HumanRange).tid], (pierCluster->HumanRange).pos, (pierCluster->HumanRange).endpos);
fprintf(fs,"VirRange=%s:%d-%d\n", h->target_name[(pierCluster->VirusRange).tid], (pierCluster->VirusRange).pos, (pierCluster->VirusRange).endpos);
for (size_t i=0; i<kv_size(pierCluster->Reads);++i) {
bam1_t *bi = kv_A(pierCluster->Reads, i);
if (sam_format1(h, bi, &ks1) < 0) {
fprintf(stderr, "Error writing output.\n");
exit_code = 1;
break;
} else {
fprintf(fs,"%s\n",ks1.s);
}
}
fprintf(fs,"\n");
//printf("HumRange=%s:%d-%d\n", h->target_name[(pierCluster->HumanRange).tid], (pierCluster->HumanRange).pos, (pierCluster->HumanRange).endpos);
//fflush(fs);
sam_plp_dectroy(pierCluster);
pierCluster = sam_plp_init();
}
}
}
/*char *qname = bam_get_qname(b);
if (sam_write1(out, h, b) < 0) {
fprintf(stderr, "[x]Error writing output.\n");
exit_code = 1;
break;
}*/
}
/* r = sam_close(out); // stdout can only be closed once
if (r < 0) {
fprintf(stderr, "Error closing output.\n");
exit_code = 1;
}
*/
hts_idx_destroy(idx);
bam_destroy1(b);
bam_destroy1(d);
bam_destroy1(d2);
bam_hdr_destroy(h);
r = sam_close(in);
free(ChrIsHum);
#ifdef DEBUGa
fflush(NULL);
//pressAnyKey();
#endif
sam_plp_dectroy(pierCluster);
//printf("<[%d]:\n",bami);
}
}
fclose(fs);
getPairedSam(NULL, NULL, NULL, NULL); // sam_close(fp2);
//printf("---[%d]---\n",exit_code);
bam_destroy1(bR1); bam_destroy1(bR2); bam_destroy1(bR3);
ks_release(&ks1);
ks_release(&ks2);
ks_release(&ks3);
return exit_code;
}