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;
}
#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 {
- int max_mq, min_mq, flag, min_baseQ, capQ_thres, max_depth;
- int openQ, extQ, tandemQ;
- char *reg, *fn_pos, *pl_list;
+ 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;
- kh_64_t *hash;
+ void *bed, *rghash;
} mplp_conf_t;
typedef struct {
bamFile fp;
bam_iter_t iter;
- int min_mq, flag, ref_id, capQ_thres;
+ bam_header_t *h;
+ int ref_id;
char *ref;
+ const mplp_conf_t *conf;
} mplp_aux_t;
typedef struct {
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->flag&MPLP_REALN)) bam_prob_realn_core(b, ma->ref, 1);
- if (has_ref && ma->capQ_thres > 10) {
- int q = bam_cap_mapQ(b, ma->ref, ma->capQ_thres);
+ 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.flag&BAM_FUNMAP) skip = 1;
- else if (b->core.qual < ma->min_mq) skip = 1;
- else if ((ma->flag&MPLP_NO_ORPHAN) && (b->core.flag&1) && !(b->core.flag&2)) skip = 1;
+ }
+ 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;
}
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);
- assert(id >= 0 && id < m->n);
+ 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]);
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, max_depth;
+ 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;
- khash_t(64) *hash = 0;
void *rghash = 0;
bcf_callaux_t *bca = 0;
for (i = 0; i < n; ++i) {
bam_header_t *h_tmp;
data[i] = calloc(1, sizeof(mplp_aux_t));
- data[i]->min_mq = conf->min_mq;
- data[i]->flag = conf->flag;
- data[i]->capQ_thres = conf->capQ_thres;
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) {
gplp.plp = calloc(sm->n, sizeof(void*));
fprintf(stderr, "[%s] %d samples in %d input files\n", __func__, sm->n, n);
- if (conf->fn_pos) hash = load_pos(conf->fn_pos, h);
// write the VCF header
if (conf->flag & MPLP_GLF) {
kstring_t s;
bh->l_smpl = s.l;
bh->sname = malloc(s.l);
memcpy(bh->sname, s.s, s.l);
- bh->l_txt = 0;
+ 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);
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);
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-sample depth to %d\n", __func__, max_depth);
+ 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 (hash) {
- khint_t k;
- k = kh_get(64, hash, (uint64_t)tid<<32 | pos);
- if (k == kh_end(hash)) continue;
- }
+ if (conf->bed && tid >= 0 && !bed_overlap(conf->bed, h->target_name[tid], pos, pos+1)) continue;
if (tid != ref_tid) {
free(ref); ref = 0;
if (conf->fai) ref = fai_fetch(conf->fai, h->target_name[tid], &ref_len);
ref_tid = tid;
}
if (conf->flag & MPLP_GLF) {
- int _ref0, ref16;
+ 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];
bcf_write(bp, bh, b);
bcf_destroy(b);
// call indels
- if (!(conf->flag&MPLP_NO_INDEL) && bcf_call_gap_prep(gplp.n, gplp.n_plp, gplp.plp, pos, bca, ref, rghash) >= 0) {
+ 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) {
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);
- if (hash) { // free the hash table
- khint_t k;
- for (k = kh_begin(hash); k < kh_end(hash); ++k)
- if (kh_exist(hash, k)) free(kh_val(hash, k));
- kh_destroy(64, hash);
- }
bcf_hdr_destroy(bh); bcf_call_destroy(bca); free(bc.PL); free(bcr);
bam_mplp_destroy(iter);
bam_header_destroy(h);
}
#define MAX_PATH_LEN 1024
-int read_file_list(const char *file_list,int *n,char **argv[])
+static int read_file_list(const char *file_list,int *n,char **argv[])
{
char buf[MAX_PATH_LEN];
int len, nfiles;
int c;
const char *file_list = NULL;
char **fn = NULL;
- int nfiles = 0;
+ int nfiles = 0, use_orphan = 0;
mplp_conf_t mplp;
memset(&mplp, 0, sizeof(mplp_conf_t));
mplp.max_mq = 60;
mplp.min_baseQ = 13;
mplp.capQ_thres = 0;
- mplp.max_depth = 250;
+ 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, "gf:r:l:M:q:Q:uaORC:BDSd:b:P:o:e:h:I")) >= 0) {
+ 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);
break;
case 'd': mplp.max_depth = atoi(optarg); break;
case 'r': mplp.reg = strdup(optarg); break;
- case 'l': mplp.fn_pos = 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 & ~MPLP_NO_ORPHAN; break;
- case 'O': mplp.flag |= MPLP_NO_ORPHAN; 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 '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, "\n");
- fprintf(stderr, "Usage: samtools mpileup [options] in1.bam [in2.bam [...]]\n\n");
- fprintf(stderr, "Options: -f FILE reference sequence file [null]\n");
- fprintf(stderr, " -r STR region in which pileup is generated [null]\n");
- fprintf(stderr, " -l FILE list of positions (format: chr pos) [null]\n");
- fprintf(stderr, " -b FILE list of input BAM files [null]\n");
- fprintf(stderr, " -M INT cap mapping quality at INT [%d]\n", mplp.max_mq);
- fprintf(stderr, " -Q INT min base quality [%d]\n", mplp.min_baseQ);
- fprintf(stderr, " -q INT filter out alignment with MQ smaller than INT [%d]\n", mplp.min_mq);
- fprintf(stderr, " -d INT max per-sample depth [%d]\n", mplp.max_depth);
- fprintf(stderr, " -P STR comma separated list of platforms for indels [all]\n");
- fprintf(stderr, " -o INT Phred-scaled gap open sequencing error probability [%d]\n", mplp.openQ);
- fprintf(stderr, " -e INT Phred-scaled gap extension seq error probability [%d]\n", mplp.extQ);
- fprintf(stderr, " -h INT coefficient for homopolyer errors [%d]\n", mplp.tandemQ);
- fprintf(stderr, " -g generate BCF output\n");
- fprintf(stderr, " -u do not compress BCF output\n");
- fprintf(stderr, " -B disable BAQ computation\n");
- fprintf(stderr, " -D output per-sample DP\n");
- fprintf(stderr, " -S output per-sample SP (strand bias P-value, slow)\n");
- fprintf(stderr, " -I do not perform indel calling\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;
}
- if ( file_list )
- {
+ 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);
+ } 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