#include <stdio.h>
#include <unistd.h>
#include <ctype.h>
+#include <string.h>
+#include <errno.h>
#include "sam.h"
#include "faidx.h"
#include "bam_maqcns.h"
#define BAM_PLF_1STBASE 0x80
#define BAM_PLF_ALLBASE 0x100
#define BAM_PLF_READPOS 0x200
+#define BAM_PLF_NOBAQ 0x400
typedef struct {
bam_header_t *h;
uint32_t format;
int tid, len, last_pos;
int mask;
+ int capQ_thres, min_baseQ;
int max_depth; // for indel calling, ignore reads with the depth too high. 0 for unlimited
char *ref;
glfFile fp_glf; // for glf output only
return hash;
}
+static inline int printw(int c, FILE *fp)
+{
+ char buf[16];
+ int l, x;
+ if (c == 0) return fputc('0', fp);
+ for (l = 0, x = c < 0? -c : c; x > 0; x /= 10) buf[l++] = x%10 + '0';
+ if (c < 0) buf[l++] = '-';
+ buf[l] = 0;
+ for (x = 0; x < l/2; ++x) {
+ int y = buf[x]; buf[x] = buf[l-1-x]; buf[l-1-x] = y;
+ }
+ fputs(buf, fp);
+ return 0;
+}
+
// an analogy to pileup_func() below
static int glt3_func(uint32_t tid, uint32_t pos, int n, const bam_pileup1_t *pu, void *data)
{
return 0;
}
-static void pileup_seq(const bam_pileup1_t *p, int pos, int ref_len, const char *ref)
+static inline void pileup_seq(const bam_pileup1_t *p, int pos, int ref_len, const char *ref)
{
- if (p->is_head) printf("^%c", p->b->core.qual > 93? 126 : p->b->core.qual + 33);
+ int j;
+ if (p->is_head) {
+ putchar('^');
+ putchar(p->b->core.qual > 93? 126 : p->b->core.qual + 33);
+ }
if (!p->is_del) {
- int j, rb, c = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos)];
- rb = (ref && pos < ref_len)? ref[pos] : 'N';
- if (c == '=' || toupper(c) == toupper(rb)) c = bam1_strand(p->b)? ',' : '.';
- else c = bam1_strand(p->b)? tolower(c) : toupper(c);
+ int c = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos)];
+ if (ref) {
+ int rb = pos < ref_len? ref[pos] : 'N';
+ if (c == '=' || bam_nt16_table[c] == bam_nt16_table[rb]) c = bam1_strand(p->b)? ',' : '.';
+ else c = bam1_strand(p->b)? tolower(c) : toupper(c);
+ } else {
+ if (c == '=') c = bam1_strand(p->b)? ',' : '.';
+ else c = bam1_strand(p->b)? tolower(c) : toupper(c);
+ }
putchar(c);
- if (p->indel > 0) {
- printf("+%d", p->indel);
- for (j = 1; j <= p->indel; ++j) {
- c = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos + j)];
- putchar(bam1_strand(p->b)? tolower(c) : toupper(c));
- }
- } else if (p->indel < 0) {
- printf("%d", p->indel);
- for (j = 1; j <= -p->indel; ++j) {
- c = (ref && (int)pos+j < ref_len)? ref[pos+j] : 'N';
- putchar(bam1_strand(p->b)? tolower(c) : toupper(c));
- }
+ } else putchar(p->is_refskip? (bam1_strand(p->b)? '<' : '>') : '*');
+ if (p->indel > 0) {
+ putchar('+'); printw(p->indel, stdout);
+ for (j = 1; j <= p->indel; ++j) {
+ int c = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos + j)];
+ putchar(bam1_strand(p->b)? tolower(c) : toupper(c));
}
- } else putchar('*');
+ } else if (p->indel < 0) {
+ printw(p->indel, stdout);
+ for (j = 1; j <= -p->indel; ++j) {
+ int c = (ref && (int)pos+j < ref_len)? ref[pos+j] : 'N';
+ putchar(bam1_strand(p->b)? tolower(c) : toupper(c));
+ }
+ }
if (p->is_tail) putchar('$');
}
}
rmsQ = (uint64_t)(sqrt((double)rmsQ / n) + .499);
cns = b<<28 | 0xf<<24 | rmsQ<<16 | 60<<8;
- } else cns = bam_maqcns_call(n, pu, d->c);
+ } else {
+ glf1_t *g = bam_maqcns_glfgen(n, pu, bam_nt16_table[rb], d->c);
+ cns = g->depth == 0? (0xfu<<28 | 0xf<<24) : glf2cns(g, (int)(d->c->q_r + .499));
+ free(g);
+ }
}
if ((d->format & (BAM_PLF_CNS|BAM_PLF_INDEL_ONLY)) && d->ref && pos < d->len) { // call indels
int m = (!d->max_depth || d->max_depth>n) ? n : d->max_depth;
}
// when only variant sites are asked for, test if the site is a variant
if ((d->format & BAM_PLF_CNS) && (d->format & BAM_PLF_VAR_ONLY)) {
- if (!(bam_nt16_table[rb] != 15 && cns>>28 != bam_nt16_table[rb])) { // not a SNP
+ if (!(bam_nt16_table[rb] != 15 && cns>>28 != 15 && cns>>28 != bam_nt16_table[rb])) { // not a SNP
if (!(r && (r->gt == 2 || strcmp(r->s[r->gt], "*")))) { // not an indel
if (r) bam_maqindel_ret_destroy(r);
return 0;
}
}
// print the first 3 columns
- printf("%s\t%d\t%c\t", d->h->target_name[tid], pos + 1, rb);
+ fputs(d->h->target_name[tid], stdout); putchar('\t');
+ printw(pos+1, stdout); putchar('\t'); putchar(rb); putchar('\t');
// print consensus information if required
if (d->format & BAM_PLF_CNS) {
- int ref_q, rb4 = bam_nt16_table[rb];
- ref_q = 0;
- if (rb4 != 15 && cns>>28 != 15 && cns>>28 != rb4) { // a SNP
- ref_q = ((cns>>24&0xf) == rb4)? cns>>8&0xff : (cns>>8&0xff) + (cns&0xff);
- if (ref_q > 255) ref_q = 255;
- }
- rms_mapq = cns>>16&0xff;
- printf("%c\t%d\t%d\t%d\t", bam_nt16_rev_table[cns>>28], cns>>8&0xff, ref_q, rms_mapq);
+ putchar(bam_nt16_rev_table[cns>>28]); putchar('\t');
+ printw(cns>>8&0xff, stdout); putchar('\t');
+ printw(cns&0xff, stdout); putchar('\t');
+ printw(cns>>16&0xff, stdout); putchar('\t');
}
// print pileup sequences
- printf("%d\t", n);
- rms_aux = 0; // we need to recalculate rms_mapq when -c is not flagged on the command line
- for (i = 0; i < n; ++i) {
- const bam_pileup1_t *p = pu + i;
- int tmp = p->b->core.qual < d->c->cap_mapQ? p->b->core.qual : d->c->cap_mapQ;
- rms_aux += tmp * tmp;
- pileup_seq(p, pos, d->len, d->ref);
- }
+ printw(n, stdout); putchar('\t');
+ for (i = 0; i < n; ++i)
+ pileup_seq(pu + i, pos, d->len, d->ref);
// finalize rms_mapq
- rms_aux = (uint64_t)(sqrt((double)rms_aux / n) + .499);
- if (rms_mapq < 0) rms_mapq = rms_aux;
+ if (d->format & BAM_PLF_CNS) {
+ for (i = rms_aux = 0; i < n; ++i) {
+ const bam_pileup1_t *p = pu + i;
+ int tmp = p->b->core.qual < d->c->cap_mapQ? p->b->core.qual : d->c->cap_mapQ;
+ rms_aux += tmp * tmp;
+ }
+ rms_aux = (uint64_t)(sqrt((double)rms_aux / n) + .499);
+ if (rms_mapq < 0) rms_mapq = rms_aux;
+ }
putchar('\t');
// print quality
for (i = 0; i < n; ++i) {
for (i = 0; i < n; ++i) {
int x = pu[i].qpos;
int l = pu[i].b->core.l_qseq;
- printf("%d,", x < l/2? x+1 : -((l-1)-x+1));
+ printw(x < l/2? x+1 : -((l-1)-x+1), stdout); putchar(',');
}
}
putchar('\n');
int c, is_SAM = 0;
char *fn_list = 0, *fn_fa = 0, *fn_pos = 0;
pu_data_t *d = (pu_data_t*)calloc(1, sizeof(pu_data_t));
- d->max_depth = 0;
- d->tid = -1; d->mask = BAM_DEF_MASK;
+ d->max_depth = 1024; d->tid = -1; d->mask = BAM_DEF_MASK; d->min_baseQ = 13;
d->c = bam_maqcns_init();
- d->c->is_soap = 1; // change the default model
+ d->c->errmod = BAM_ERRMOD_MAQ2; // change the default model
d->ido = bam_maqindel_opt_init();
- while ((c = getopt(argc, argv, "st:f:cT:N:r:l:d:im:gI:G:vM:S2aR:PA")) >= 0) {
+ while ((c = getopt(argc, argv, "st:f:cT:N:r:l:d:im:gI:G:vM:S2aR:PAQ:C:B")) >= 0) {
switch (c) {
- case 'a': d->c->is_soap = 1; break;
- case 'A': d->c->is_soap = 0; break;
+ case 'Q': d->c->min_baseQ = atoi(optarg); break;
+ case 'C': d->capQ_thres = atoi(optarg); break;
+ case 'B': d->format |= BAM_PLF_NOBAQ; break;
+ case 'a': d->c->errmod = BAM_ERRMOD_SOAP; break;
+ case 'A': d->c->errmod = BAM_ERRMOD_MAQ; break;
case 's': d->format |= BAM_PLF_SIMPLE; break;
case 't': fn_list = strdup(optarg); break;
case 'l': fn_pos = strdup(optarg); break;
default: fprintf(stderr, "Unrecognizd option '-%c'.\n", c); return 1;
}
}
+ if (d->c->errmod != BAM_ERRMOD_MAQ2) d->c->theta += 0.02;
+ if (d->c->theta > 1.0) d->c->theta = 1.0;
if (fn_list) is_SAM = 1;
if (optind == argc) {
fprintf(stderr, "\n");
fprintf(stderr, "Usage: samtools pileup [options] <in.bam>|<in.sam>\n\n");
fprintf(stderr, "Option: -s simple (yet incomplete) pileup format\n");
fprintf(stderr, " -S the input is in SAM\n");
- fprintf(stderr, " -A use the MAQ model for SNP calling\n");
+ fprintf(stderr, " -B disable BAQ computation\n");
+ fprintf(stderr, " -A use the original MAQ model for SNP calling (DEPRECATED)\n");
fprintf(stderr, " -2 output the 2nd best call and quality\n");
fprintf(stderr, " -i only show lines/consensus with indels\n");
- fprintf(stderr, " -m INT filtering reads with bits in INT [%d]\n", d->mask);
+ fprintf(stderr, " -Q INT min base quality (possibly capped by BAQ) [%d]\n", d->c->min_baseQ);
+ fprintf(stderr, " -C INT coefficient for adjusting mapQ of poor mappings [%d]\n", d->capQ_thres);
+ fprintf(stderr, " -m INT filtering reads with bits in INT [0x%x]\n", d->mask);
fprintf(stderr, " -M INT cap mapping quality at INT [%d]\n", d->c->cap_mapQ);
- fprintf(stderr, " -d INT limit maximum depth for indels [unlimited]\n");
+ fprintf(stderr, " -d INT limit maximum depth for indels [%d]\n", d->max_depth);
fprintf(stderr, " -t FILE list of reference sequences (force -S)\n");
fprintf(stderr, " -l FILE list of sites at which pileup is output\n");
fprintf(stderr, " -f FILE reference sequence in the FASTA format\n\n");
- fprintf(stderr, " -c output the SOAPsnp consensus sequence\n");
+ fprintf(stderr, " -c compute the consensus sequence\n");
fprintf(stderr, " -v print variants only (for -c)\n");
- fprintf(stderr, " -g output in the GLFv3 format (suppressing -c/-i/-s)\n");
- fprintf(stderr, " -T FLOAT theta in maq consensus calling model (for -c/-g) [%f]\n", d->c->theta);
- fprintf(stderr, " -N INT number of haplotypes in the sample (for -c/-g) [%d]\n", d->c->n_hap);
- fprintf(stderr, " -r FLOAT prior of a difference between two haplotypes (for -c/-g) [%f]\n", d->c->het_rate);
- fprintf(stderr, " -G FLOAT prior of an indel between two haplotypes (for -c/-g) [%f]\n", d->ido->r_indel);
- fprintf(stderr, " -I INT phred prob. of an indel in sequencing/prep. (for -c/-g) [%d]\n", d->ido->q_indel);
+ fprintf(stderr, " -g output in the GLFv3 format (DEPRECATED)\n");
+ fprintf(stderr, " -T FLOAT theta in maq consensus calling model (for -c) [%.4g]\n", d->c->theta);
+ fprintf(stderr, " -N INT number of haplotypes in the sample (for -c) [%d]\n", d->c->n_hap);
+ fprintf(stderr, " -r FLOAT prior of a difference between two haplotypes (for -c) [%.4g]\n", d->c->het_rate);
+ fprintf(stderr, " -G FLOAT prior of an indel between two haplotypes (for -c) [%.4g]\n", d->ido->r_indel);
+ fprintf(stderr, " -I INT phred prob. of an indel in sequencing/prep. (for -c) [%d]\n", d->ido->q_indel);
fprintf(stderr, "\n");
+ fprintf(stderr, "Warning: Please use the `mpileup' command instead `pileup'. `Pileup' is deprecated!\n\n");
free(fn_list); free(fn_fa); free(d);
return 1;
}
}
d->h = fp->header;
if (fn_pos) d->hash = load_pos(fn_pos, d->h);
- sampileup(fp, d->mask, pileup_func, d);
+ { // run pileup
+ extern int bam_prob_realn(bam1_t *b, const char *ref);
+ extern int bam_cap_mapQ(bam1_t *b, char *ref, int thres);
+ bam1_t *b;
+ int ret, tid, pos, n_plp;
+ bam_plp_t iter;
+ const bam_pileup1_t *plp;
+ b = bam_init1();
+ iter = bam_plp_init(0, 0);
+ bam_plp_set_mask(iter, d->mask);
+ while ((ret = samread(fp, b)) >= 0) {
+ int skip = 0;
+ if ((int)b->core.tid < 0) break;
+ // update d->ref if necessary
+ if (d->fai && (int)b->core.tid != d->tid) {
+ free(d->ref);
+ d->ref = faidx_fetch_seq(d->fai, d->h->target_name[b->core.tid], 0, 0x7fffffff, &d->len);
+ d->tid = b->core.tid;
+ }
+ if (d->ref && (d->format&BAM_PLF_CNS) && !(d->format&BAM_PLF_NOBAQ)) bam_prob_realn(b, d->ref);
+ if (d->ref && (d->format&BAM_PLF_CNS) && d->capQ_thres > 10) {
+ int q = bam_cap_mapQ(b, d->ref, d->capQ_thres);
+ if (q < 0) skip = 1;
+ else if (b->core.qual > q) b->core.qual = q;
+ } else if (b->core.flag&BAM_FUNMAP) skip = 1;
+ else if ((d->format&BAM_PLF_CNS) && (b->core.flag&1) && !(b->core.flag&2)) skip = 1;
+ if (skip) continue;
+ bam_plp_push(iter, b);
+ while ((plp = bam_plp_next(iter, &tid, &pos, &n_plp)) != 0)
+ pileup_func(tid, pos, n_plp, plp, d);
+ }
+ bam_plp_push(iter, 0);
+ while ((plp = bam_plp_next(iter, &tid, &pos, &n_plp)) != 0)
+ pileup_func(tid, pos, n_plp, plp, d);
+ bam_plp_destroy(iter);
+ bam_destroy1(b);
+ }
samclose(fp); // d->h will be destroyed here
}
* mpileup *
***********/
+#include <assert.h>
+#include "bam2bcf.h"
+#include "sample.h"
+
+#define MPLP_GLF 0x10
+#define MPLP_NO_COMP 0x20
+#define MPLP_NO_ORPHAN 0x40
+#define MPLP_REALN 0x80
+#define MPLP_FMT_DP 0x100
+#define MPLP_FMT_SP 0x200
+#define MPLP_NO_INDEL 0x400
+#define MPLP_EXT_BAQ 0x800
+#define MPLP_ILLUMINA13 0x1000
+
+void *bed_read(const char *fn);
+void bed_destroy(void *_h);
+int bed_overlap(const void *_h, const char *chr, int beg, int end);
+
typedef struct {
- char *reg;
+ int max_mq, min_mq, flag, min_baseQ, capQ_thres, max_depth, max_indel_depth;
+ int openQ, extQ, tandemQ, min_support; // for indels
+ double min_frac; // for indels
+ char *reg, *pl_list;
faidx_t *fai;
+ void *bed, *rghash;
} mplp_conf_t;
typedef struct {
bamFile fp;
bam_iter_t iter;
+ bam_header_t *h;
+ int ref_id;
+ char *ref;
+ const mplp_conf_t *conf;
} mplp_aux_t;
+typedef struct {
+ int n;
+ int *n_plp, *m_plp;
+ bam_pileup1_t **plp;
+} mplp_pileup_t;
+
static int mplp_func(void *data, bam1_t *b)
{
+ extern int bam_realn(bam1_t *b, const char *ref);
+ extern int bam_prob_realn_core(bam1_t *b, const char *ref, int);
+ extern int bam_cap_mapQ(bam1_t *b, char *ref, int thres);
mplp_aux_t *ma = (mplp_aux_t*)data;
- if (ma->iter) return bam_iter_read(ma->fp, ma->iter, b);
- return bam_read1(ma->fp, b);
+ int ret, skip = 0;
+ do {
+ int has_ref;
+ ret = ma->iter? bam_iter_read(ma->fp, ma->iter, b) : bam_read1(ma->fp, b);
+ if (ret < 0) break;
+ if (b->core.tid < 0 || (b->core.flag&BAM_FUNMAP)) { // exclude unmapped reads
+ skip = 1;
+ continue;
+ }
+ if (ma->conf->bed) { // test overlap
+ skip = !bed_overlap(ma->conf->bed, ma->h->target_name[b->core.tid], b->core.pos, bam_calend(&b->core, bam1_cigar(b)));
+ if (skip) continue;
+ }
+ if (ma->conf->rghash) { // exclude read groups
+ uint8_t *rg = bam_aux_get(b, "RG");
+ skip = (rg && bcf_str2id(ma->conf->rghash, (const char*)(rg+1)) >= 0);
+ if (skip) continue;
+ }
+ if (ma->conf->flag & MPLP_ILLUMINA13) {
+ int i;
+ uint8_t *qual = bam1_qual(b);
+ for (i = 0; i < b->core.l_qseq; ++i)
+ qual[i] = qual[i] > 31? qual[i] - 31 : 0;
+ }
+ has_ref = (ma->ref && ma->ref_id == b->core.tid)? 1 : 0;
+ skip = 0;
+ if (has_ref && (ma->conf->flag&MPLP_REALN)) bam_prob_realn_core(b, ma->ref, (ma->conf->flag & MPLP_EXT_BAQ)? 3 : 1);
+ if (has_ref && ma->conf->capQ_thres > 10) {
+ int q = bam_cap_mapQ(b, ma->ref, ma->conf->capQ_thres);
+ if (q < 0) skip = 1;
+ else if (b->core.qual > q) b->core.qual = q;
+ }
+ else if (b->core.qual < ma->conf->min_mq) skip = 1;
+ else if ((ma->conf->flag&MPLP_NO_ORPHAN) && (b->core.flag&1) && !(b->core.flag&2)) skip = 1;
+ } while (skip);
+ return ret;
+}
+
+static void group_smpl(mplp_pileup_t *m, bam_sample_t *sm, kstring_t *buf,
+ int n, char *const*fn, int *n_plp, const bam_pileup1_t **plp)
+{
+ int i, j;
+ memset(m->n_plp, 0, m->n * sizeof(int));
+ for (i = 0; i < n; ++i) {
+ for (j = 0; j < n_plp[i]; ++j) {
+ const bam_pileup1_t *p = plp[i] + j;
+ uint8_t *q;
+ int id = -1;
+ q = bam_aux_get(p->b, "RG");
+ if (q) id = bam_smpl_rg2smid(sm, fn[i], (char*)q+1, buf);
+ if (id < 0) id = bam_smpl_rg2smid(sm, fn[i], 0, buf);
+ if (id < 0 || id >= m->n) {
+ assert(q); // otherwise a bug
+ fprintf(stderr, "[%s] Read group %s used in file %s but absent from the header or an alignment missing read group.\n", __func__, (char*)q+1, fn[i]);
+ exit(1);
+ }
+ if (m->n_plp[id] == m->m_plp[id]) {
+ m->m_plp[id] = m->m_plp[id]? m->m_plp[id]<<1 : 8;
+ m->plp[id] = realloc(m->plp[id], sizeof(bam_pileup1_t) * m->m_plp[id]);
+ }
+ m->plp[id][m->n_plp[id]++] = *p;
+ }
+ }
}
static int mpileup(mplp_conf_t *conf, int n, char **fn)
{
+ extern void *bcf_call_add_rg(void *rghash, const char *hdtext, const char *list);
+ extern void bcf_call_del_rghash(void *rghash);
mplp_aux_t **data;
- int i, tid, pos, *n_plp, beg0 = 0, end0 = 1u<<29, ref_len, ref_tid;
+ int i, tid, pos, *n_plp, beg0 = 0, end0 = 1u<<29, ref_len, ref_tid, max_depth, max_indel_depth;
const bam_pileup1_t **plp;
bam_mplp_t iter;
bam_header_t *h = 0;
char *ref;
- // allocate
+ void *rghash = 0;
+
+ bcf_callaux_t *bca = 0;
+ bcf_callret1_t *bcr = 0;
+ bcf_call_t bc;
+ bcf_t *bp = 0;
+ bcf_hdr_t *bh = 0;
+
+ bam_sample_t *sm = 0;
+ kstring_t buf;
+ mplp_pileup_t gplp;
+
+ memset(&gplp, 0, sizeof(mplp_pileup_t));
+ memset(&buf, 0, sizeof(kstring_t));
+ memset(&bc, 0, sizeof(bcf_call_t));
data = calloc(n, sizeof(void*));
plp = calloc(n, sizeof(void*));
n_plp = calloc(n, sizeof(int*));
+ sm = bam_smpl_init();
+
// read the header and initialize data
for (i = 0; i < n; ++i) {
bam_header_t *h_tmp;
data[i] = calloc(1, sizeof(mplp_aux_t));
- data[i]->fp = bam_open(fn[i], "r");
+ data[i]->fp = strcmp(fn[i], "-") == 0? bam_dopen(fileno(stdin), "r") : bam_open(fn[i], "r");
+ data[i]->conf = conf;
h_tmp = bam_header_read(data[i]->fp);
+ data[i]->h = i? h : h_tmp; // for i==0, "h" has not been set yet
+ bam_smpl_add(sm, fn[i], h_tmp->text);
+ rghash = bcf_call_add_rg(rghash, h_tmp->text, conf->pl_list);
if (conf->reg) {
int beg, end;
bam_index_t *idx;
bam_header_destroy(h_tmp);
}
}
- // mpileup
+ gplp.n = sm->n;
+ gplp.n_plp = calloc(sm->n, sizeof(int));
+ gplp.m_plp = calloc(sm->n, sizeof(int));
+ gplp.plp = calloc(sm->n, sizeof(void*));
+
+ fprintf(stderr, "[%s] %d samples in %d input files\n", __func__, sm->n, n);
+ // write the VCF header
+ if (conf->flag & MPLP_GLF) {
+ kstring_t s;
+ bh = calloc(1, sizeof(bcf_hdr_t));
+ s.l = s.m = 0; s.s = 0;
+ bp = bcf_open("-", (conf->flag&MPLP_NO_COMP)? "wu" : "w");
+ for (i = 0; i < h->n_targets; ++i) {
+ kputs(h->target_name[i], &s);
+ kputc('\0', &s);
+ }
+ bh->l_nm = s.l;
+ bh->name = malloc(s.l);
+ memcpy(bh->name, s.s, s.l);
+ s.l = 0;
+ for (i = 0; i < sm->n; ++i) {
+ kputs(sm->smpl[i], &s); kputc('\0', &s);
+ }
+ bh->l_smpl = s.l;
+ bh->sname = malloc(s.l);
+ memcpy(bh->sname, s.s, s.l);
+ bh->txt = malloc(strlen(BAM_VERSION) + 64);
+ bh->l_txt = 1 + sprintf(bh->txt, "##samtoolsVersion=%s\n", BAM_VERSION);
+ free(s.s);
+ bcf_hdr_sync(bh);
+ bcf_hdr_write(bp, bh);
+ bca = bcf_call_init(-1., conf->min_baseQ);
+ bcr = calloc(sm->n, sizeof(bcf_callret1_t));
+ bca->rghash = rghash;
+ bca->openQ = conf->openQ, bca->extQ = conf->extQ, bca->tandemQ = conf->tandemQ;
+ bca->min_frac = conf->min_frac;
+ bca->min_support = conf->min_support;
+ }
ref_tid = -1; ref = 0;
iter = bam_mplp_init(n, mplp_func, (void**)data);
+ max_depth = conf->max_depth;
+ if (max_depth * sm->n > 1<<20)
+ fprintf(stderr, "(%s) Max depth is above 1M. Potential memory hog!\n", __func__);
+ if (max_depth * sm->n < 8000) {
+ max_depth = 8000 / sm->n;
+ fprintf(stderr, "<%s> Set max per-file depth to %d\n", __func__, max_depth);
+ }
+ max_indel_depth = conf->max_indel_depth * sm->n;
+ bam_mplp_set_maxcnt(iter, max_depth);
while (bam_mplp_auto(iter, &tid, &pos, n_plp, plp) > 0) {
if (conf->reg && (pos < beg0 || pos >= end0)) continue; // out of the region requested
+ if (conf->bed && tid >= 0 && !bed_overlap(conf->bed, h->target_name[tid], pos, pos+1)) continue;
if (tid != ref_tid) {
- free(ref);
+ free(ref); ref = 0;
if (conf->fai) ref = fai_fetch(conf->fai, h->target_name[tid], &ref_len);
+ for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid;
ref_tid = tid;
}
- printf("%s\t%d\t%c", h->target_name[tid], pos + 1, (ref && pos < ref_len)? ref[pos] : 'N');
- for (i = 0; i < n; ++i) {
- int j;
- printf("\t%d\t", n_plp[i]);
- if (n_plp[i] == 0) printf("*\t*");
- else {
- for (j = 0; j < n_plp[i]; ++j)
- pileup_seq(plp[i] + j, pos, ref_len, ref);
- putchar('\t');
- for (j = 0; j < n_plp[i]; ++j) {
- const bam_pileup1_t *p = plp[i] + j;
- int c = bam1_qual(p->b)[p->qpos] + 33;
- if (c > 126) c = 126;
- putchar(c);
+ if (conf->flag & MPLP_GLF) {
+ int total_depth, _ref0, ref16;
+ bcf1_t *b = calloc(1, sizeof(bcf1_t));
+ for (i = total_depth = 0; i < n; ++i) total_depth += n_plp[i];
+ group_smpl(&gplp, sm, &buf, n, fn, n_plp, plp);
+ _ref0 = (ref && pos < ref_len)? ref[pos] : 'N';
+ ref16 = bam_nt16_table[_ref0];
+ for (i = 0; i < gplp.n; ++i)
+ bcf_call_glfgen(gplp.n_plp[i], gplp.plp[i], ref16, bca, bcr + i);
+ bcf_call_combine(gplp.n, bcr, ref16, &bc);
+ bcf_call2bcf(tid, pos, &bc, b, (conf->flag&(MPLP_FMT_DP|MPLP_FMT_SP))? bcr : 0,
+ (conf->flag&MPLP_FMT_SP), 0, 0);
+ bcf_write(bp, bh, b);
+ bcf_destroy(b);
+ // call indels
+ if (!(conf->flag&MPLP_NO_INDEL) && total_depth < max_indel_depth && bcf_call_gap_prep(gplp.n, gplp.n_plp, gplp.plp, pos, bca, ref, rghash) >= 0) {
+ for (i = 0; i < gplp.n; ++i)
+ bcf_call_glfgen(gplp.n_plp[i], gplp.plp[i], -1, bca, bcr + i);
+ if (bcf_call_combine(gplp.n, bcr, -1, &bc) >= 0) {
+ b = calloc(1, sizeof(bcf1_t));
+ bcf_call2bcf(tid, pos, &bc, b, (conf->flag&(MPLP_FMT_DP|MPLP_FMT_SP))? bcr : 0,
+ (conf->flag&MPLP_FMT_SP), bca, ref);
+ bcf_write(bp, bh, b);
+ bcf_destroy(b);
}
}
+ } else {
+ printf("%s\t%d\t%c", h->target_name[tid], pos + 1, (ref && pos < ref_len)? ref[pos] : 'N');
+ for (i = 0; i < n; ++i) {
+ int j;
+ printf("\t%d\t", n_plp[i]);
+ if (n_plp[i] == 0) printf("*\t*");
+ else {
+ for (j = 0; j < n_plp[i]; ++j)
+ pileup_seq(plp[i] + j, pos, ref_len, ref);
+ putchar('\t');
+ for (j = 0; j < n_plp[i]; ++j) {
+ const bam_pileup1_t *p = plp[i] + j;
+ int c = bam1_qual(p->b)[p->qpos] + 33;
+ if (c > 126) c = 126;
+ putchar(c);
+ }
+ }
+ }
+ putchar('\n');
}
- putchar('\n');
}
+
+ bcf_close(bp);
+ bam_smpl_destroy(sm); free(buf.s);
+ for (i = 0; i < gplp.n; ++i) free(gplp.plp[i]);
+ free(gplp.plp); free(gplp.n_plp); free(gplp.m_plp);
+ bcf_call_del_rghash(rghash);
+ bcf_hdr_destroy(bh); bcf_call_destroy(bca); free(bc.PL); free(bcr);
bam_mplp_destroy(iter);
bam_header_destroy(h);
for (i = 0; i < n; ++i) {
return 0;
}
+#define MAX_PATH_LEN 1024
+static int read_file_list(const char *file_list,int *n,char **argv[])
+{
+ char buf[MAX_PATH_LEN];
+ int len, nfiles;
+ char **files;
+
+ FILE *fh = fopen(file_list,"r");
+ if ( !fh )
+ {
+ fprintf(stderr,"%s: %s\n", file_list,strerror(errno));
+ return 1;
+ }
+
+ // Speed is not an issue here, determine the number of files by reading the file twice
+ nfiles = 0;
+ while ( fgets(buf,MAX_PATH_LEN,fh) ) nfiles++;
+
+ if ( fseek(fh, 0L, SEEK_SET) )
+ {
+ fprintf(stderr,"%s: %s\n", file_list,strerror(errno));
+ return 1;
+ }
+
+ files = calloc(nfiles,sizeof(char*));
+ nfiles = 0;
+ while ( fgets(buf,MAX_PATH_LEN,fh) )
+ {
+ len = strlen(buf);
+ while ( len>0 && isspace(buf[len-1]) ) len--;
+ if ( !len ) continue;
+
+ files[nfiles] = malloc(sizeof(char)*(len+1));
+ strncpy(files[nfiles],buf,len);
+ files[nfiles][len] = 0;
+ nfiles++;
+ }
+ fclose(fh);
+ if ( !nfiles )
+ {
+ fprintf(stderr,"No files read from %s\n", file_list);
+ return 1;
+ }
+ *argv = files;
+ *n = nfiles;
+ return 0;
+}
+#undef MAX_PATH_LEN
+
int bam_mpileup(int argc, char *argv[])
{
int c;
+ const char *file_list = NULL;
+ char **fn = NULL;
+ int nfiles = 0, use_orphan = 0;
mplp_conf_t mplp;
memset(&mplp, 0, sizeof(mplp_conf_t));
- while ((c = getopt(argc, argv, "f:r:")) >= 0) {
+ mplp.max_mq = 60;
+ mplp.min_baseQ = 13;
+ mplp.capQ_thres = 0;
+ mplp.max_depth = 250; mplp.max_indel_depth = 250;
+ mplp.openQ = 40; mplp.extQ = 20; mplp.tandemQ = 100;
+ mplp.min_frac = 0.002; mplp.min_support = 1;
+ mplp.flag = MPLP_NO_ORPHAN | MPLP_REALN;
+ while ((c = getopt(argc, argv, "Agf:r:l:M:q:Q:uaRC:BDSd:L:b:P:o:e:h:Im:F:EG:6")) >= 0) {
switch (c) {
case 'f':
mplp.fai = fai_load(optarg);
if (mplp.fai == 0) return 1;
break;
- case 'r': mplp.reg = strdup(optarg);
+ case 'd': mplp.max_depth = atoi(optarg); break;
+ case 'r': mplp.reg = strdup(optarg); break;
+ case 'l': mplp.bed = bed_read(optarg); break;
+ case 'P': mplp.pl_list = strdup(optarg); break;
+ case 'g': mplp.flag |= MPLP_GLF; break;
+ case 'u': mplp.flag |= MPLP_NO_COMP | MPLP_GLF; break;
+ case 'a': mplp.flag |= MPLP_NO_ORPHAN | MPLP_REALN; break;
+ case 'B': mplp.flag &= ~MPLP_REALN; break;
+ case 'R': mplp.flag |= MPLP_REALN; break;
+ case 'D': mplp.flag |= MPLP_FMT_DP; break;
+ case 'S': mplp.flag |= MPLP_FMT_SP; break;
+ case 'I': mplp.flag |= MPLP_NO_INDEL; break;
+ case 'E': mplp.flag |= MPLP_EXT_BAQ; break;
+ case '6': mplp.flag |= MPLP_ILLUMINA13; break;
+ case 'C': mplp.capQ_thres = atoi(optarg); break;
+ case 'M': mplp.max_mq = atoi(optarg); break;
+ case 'q': mplp.min_mq = atoi(optarg); break;
+ case 'Q': mplp.min_baseQ = atoi(optarg); break;
+ case 'b': file_list = optarg; break;
+ case 'o': mplp.openQ = atoi(optarg); break;
+ case 'e': mplp.extQ = atoi(optarg); break;
+ case 'h': mplp.tandemQ = atoi(optarg); break;
+ case 'A': use_orphan = 1; break;
+ case 'F': mplp.min_frac = atof(optarg); break;
+ case 'm': mplp.min_support = atoi(optarg); break;
+ case 'L': mplp.max_indel_depth = atoi(optarg); break;
+ case 'G': {
+ FILE *fp_rg;
+ char buf[1024];
+ mplp.rghash = bcf_str2id_init();
+ if ((fp_rg = fopen(optarg, "r")) == 0)
+ fprintf(stderr, "(%s) Fail to open file %s. Continue anyway.\n", __func__, optarg);
+ while (!feof(fp_rg) && fscanf(fp_rg, "%s", buf) > 0) // this is not a good style, but forgive me...
+ bcf_str2id_add(mplp.rghash, strdup(buf));
+ fclose(fp_rg);
+ }
+ break;
}
}
+ if (use_orphan) mplp.flag &= ~MPLP_NO_ORPHAN;
if (argc == 1) {
- fprintf(stderr, "Usage: samtools mpileup [-r reg] [-f in.fa] in1.bam [in2.bam [...]]\n");
+ fprintf(stderr, "\n");
+ fprintf(stderr, "Usage: samtools mpileup [options] in1.bam [in2.bam [...]]\n\n");
+ fprintf(stderr, "Input options:\n\n");
+ fprintf(stderr, " -6 assume the quality is in the Illumina-1.3+ encoding\n");
+ fprintf(stderr, " -A count anomalous read pairs\n");
+ fprintf(stderr, " -B disable BAQ computation\n");
+ fprintf(stderr, " -b FILE list of input BAM files [null]\n");
+ fprintf(stderr, " -d INT max per-BAM depth to avoid excessive memory usage [%d]\n", mplp.max_depth);
+ fprintf(stderr, " -E extended BAQ for higher sensitivity but lower specificity\n");
+ fprintf(stderr, " -f FILE faidx indexed reference sequence file [null]\n");
+ fprintf(stderr, " -G FILE exclude read groups listed in FILE [null]\n");
+ fprintf(stderr, " -l FILE list of positions (chr pos) or regions (BED) [null]\n");
+ fprintf(stderr, " -M INT cap mapping quality at INT [%d]\n", mplp.max_mq);
+ fprintf(stderr, " -r STR region in which pileup is generated [null]\n");
+ fprintf(stderr, " -q INT skip alignments with mapQ smaller than INT [%d]\n", mplp.min_mq);
+ fprintf(stderr, " -Q INT skip bases with baseQ/BAQ smaller than INT [%d]\n", mplp.min_baseQ);
+ fprintf(stderr, "\nOutput options:\n\n");
+ fprintf(stderr, " -D output per-sample DP in BCF (require -g/-u)\n");
+ fprintf(stderr, " -g generate BCF output (genotype likelihoods)\n");
+ fprintf(stderr, " -S output per-sample strand bias P-value in BCF (require -g/-u)\n");
+ fprintf(stderr, " -u generate uncompress BCF output\n");
+ fprintf(stderr, "\nSNP/INDEL genotype likelihoods options (effective with `-g' or `-u'):\n\n");
+ fprintf(stderr, " -e INT Phred-scaled gap extension seq error probability [%d]\n", mplp.extQ);
+ fprintf(stderr, " -F FLOAT minimum fraction of gapped reads for candidates [%g]\n", mplp.min_frac);
+ fprintf(stderr, " -h INT coefficient for homopolymer errors [%d]\n", mplp.tandemQ);
+ fprintf(stderr, " -I do not perform indel calling\n");
+ fprintf(stderr, " -L INT max per-sample depth for INDEL calling [%d]\n", mplp.max_indel_depth);
+ fprintf(stderr, " -m INT minimum gapped reads for indel candidates [%d]\n", mplp.min_support);
+ fprintf(stderr, " -o INT Phred-scaled gap open sequencing error probability [%d]\n", mplp.openQ);
+ fprintf(stderr, " -P STR comma separated list of platforms for indels [all]\n");
+ fprintf(stderr, "\n");
+ fprintf(stderr, "Notes: Assuming diploid individuals.\n\n");
return 1;
}
- mpileup(&mplp, argc - optind, argv + optind);
- free(mplp.reg);
+ if (file_list) {
+ if ( read_file_list(file_list,&nfiles,&fn) ) return 1;
+ mpileup(&mplp,nfiles,fn);
+ for (c=0; c<nfiles; c++) free(fn[c]);
+ free(fn);
+ } else mpileup(&mplp, argc - optind, argv + optind);
+ if (mplp.rghash) bcf_str2id_thorough_destroy(mplp.rghash);
+ free(mplp.reg); free(mplp.pl_list);
if (mplp.fai) fai_destroy(mplp.fai);
+ if (mplp.bed) bed_destroy(mplp.bed);
return 0;
}
projects / samtools.git / blobdiff