Esempio n. 1
0
void seekRegion(BamReaderData * data) {
	int tid, beg, end;

	if (data->conf->reg) {
		// Create BAM iterator at region
		if (bam_parse_region(data->data->h, data->conf->reg, &tid, &beg, &end) < 0) {
			fprintf(stderr, "[%s] malformatted region or wrong seqname for input.\n", __func__);
			exit(1);
		}
		data->data->iter = bam_iter_query(data->idx, tid, beg, end);
		data->ref_tid = tid;
	} else {
		// Create general BAM iterator
		data->data->iter = NULL;
	}

	// Create pileup iterator	
	data->iter = bam_mplp_init(1, mplp_func, (void**) &data->data);
}
Esempio n. 2
0
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, tid0 = -1, beg0 = 0, end0 = 1u<<29, ref_len, ref_tid = -1, max_depth, max_indel_depth;
	const bam_pileup1_t **plp;
	bam_mplp_t iter;
	bam_header_t *h = 0;
	char *ref;
	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 = 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], (conf->flag&MPLP_IGNORE_RG)? 0 : 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;
			idx = bam_index_load(fn[i]);
			if (idx == 0) {
				fprintf(stderr, "[%s] fail to load index for %d-th input.\n", __func__, i+1);
				exit(1);
			}
			if (bam_parse_region(h_tmp, conf->reg, &tid, &beg, &end) < 0) {
				fprintf(stderr, "[%s] malformatted region or wrong seqname for %d-th input.\n", __func__, i+1);
				exit(1);
			}
			if (i == 0) tid0 = tid, beg0 = beg, end0 = end;
			data[i]->iter = bam_iter_query(idx, tid, beg, end);
			bam_index_destroy(idx);
		}
		if (i == 0) h = h_tmp;
		else {
			// FIXME: to check consistency
			bam_header_destroy(h_tmp);
		}
	}
	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;
	}
	if (tid0 >= 0 && conf->fai) { // region is set
		ref = faidx_fetch_seq(conf->fai, h->target_name[tid0], 0, 0x7fffffff, &ref_len);
		ref_tid = tid0;
		for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid0;
	} else 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); ref = 0;
			if (conf->fai) ref = faidx_fetch_seq(conf->fai, h->target_name[tid], 0, 0x7fffffff, &ref_len);
			for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid;
			ref_tid = tid;
		}
		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, conf->flag & MPLP_IGNORE_RG);
			_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*"); // FIXME: printf() is very slow...
					if (conf->flag & MPLP_PRINT_POS) 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_PRINT_MAPQ) {
						putchar('\t');
						for (j = 0; j < n_plp[i]; ++j) {
							int c = plp[i][j].b->core.qual + 33;
							if (c > 126) c = 126;
							putchar(c);
						}
					}
					if (conf->flag & MPLP_PRINT_POS) {
						putchar('\t');
						for (j = 0; j < n_plp[i]; ++j) {
							if (j > 0) putchar(',');
							printf("%d", plp[i][j].qpos + 1); // FIXME: printf() is very slow...
						}
					}
				}
			}
			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) {
		bam_close(data[i]->fp);
		if (data[i]->iter) bam_iter_destroy(data[i]->iter);
		free(data[i]);
	}
	free(data); free(plp); free(ref); free(n_plp);
	return 0;
}
Esempio n. 3
0
int main_depth(int argc, char *argv[])
#endif
{
	int i, n, tid, beg, end, pos, *n_plp, baseQ = 0, mapQ = 0;
	const bam_pileup1_t **plp;
	char *reg = 0; // specified region
	void *bed = 0; // BED data structure
	bam_header_t *h = 0; // BAM header of the 1st input
	aux_t **data;
	bam_mplp_t mplp;

	// parse the command line
	while ((n = getopt(argc, argv, "r:b:q:Q:")) >= 0) {
		switch (n) {
			case 'r': reg = strdup(optarg); break;   // parsing a region requires a BAM header
			case 'b': bed = bed_read(optarg); break; // BED or position list file can be parsed now
			case 'q': baseQ = atoi(optarg); break;   // base quality threshold
			case 'Q': mapQ = atoi(optarg); break;    // mapping quality threshold
		}
	}
	if (optind == argc) {
		fprintf(stderr, "Usage: bam2depth [-r reg] [-q baseQthres] [-Q mapQthres] [-b in.bed] <in1.bam> [...]\n");
		return 1;
	}

	// initialize the auxiliary data structures
	n = argc - optind; // the number of BAMs on the command line
	data = (aux_t **) calloc(n, sizeof(void*)); // data[i] for the i-th input
	beg = 0; end = 1<<30; tid = -1;  // set the default region
	for (i = 0; i < n; ++i) {
		bam_header_t *htmp;
		data[i] = (aux_t *) calloc(1, sizeof(aux_t));
		data[i]->fp = bam_open(argv[optind+i], "r"); // open BAM
		data[i]->min_mapQ = mapQ;                    // set the mapQ filter
		htmp = bam_header_read(data[i]->fp);         // read the BAM header
		if (i == 0) {
			h = htmp; // keep the header of the 1st BAM
			if (reg) bam_parse_region(h, reg, &tid, &beg, &end); // also parse the region
		} else bam_header_destroy(htmp); // if not the 1st BAM, trash the header
		if (tid >= 0) { // if a region is specified and parsed successfully
			bam_index_t *idx = bam_index_load(argv[optind+i]);  // load the index
			data[i]->iter = bam_iter_query(idx, tid, beg, end); // set the iterator
			bam_index_destroy(idx); // the index is not needed any more; phase out of the memory
		}
	}

	// the core multi-pileup loop
	mplp = bam_mplp_init(n, read_bam, (void**)data); // initialization
	n_plp = (int*) calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM
	plp = (bam_pileup1_t **) calloc(n, sizeof(void*)); // plp[i] points to the array of covering reads (internal in mplp)
	while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0) { // come to the next covered position
		if (pos < beg || pos >= end) continue; // out of range; skip
		if (bed && bed_overlap(bed, h->target_name[tid], pos, pos + 1) == 0) continue; // not in BED; skip
		fputs(h->target_name[tid], stdout); printf("\t%d", pos+1); // a customized printf() would be faster
		for (i = 0; i < n; ++i) { // base level filters have to go here
			int j, m = 0;
			for (j = 0; j < n_plp[i]; ++j) {
				const bam_pileup1_t *p = plp[i] + j; // DON'T modfity plp[][] unless you really know
				if (p->is_del || p->is_refskip) ++m; // having dels or refskips at tid:pos
				else if (bam1_qual(p->b)[p->qpos] < baseQ) ++m; // low base quality
			}
			printf("\t%d", n_plp[i] - m); // this the depth to output
		}
		putchar('\n');
	}
	free(n_plp); free(plp);
	bam_mplp_destroy(mplp);

	bam_header_destroy(h);
	for (i = 0; i < n; ++i) {
		bam_close(data[i]->fp);
		if (data[i]->iter) bam_iter_destroy(data[i]->iter);
		free(data[i]);
	}
	free(data); free(reg);
	if (bed) bed_destroy(bed);
	return 0;
}
Esempio n. 4
0
int main_bedcov(int argc, char *argv[])
{
    gzFile fp;
    kstring_t str;
    kstream_t *ks;
    hts_idx_t **idx;
    aux_t **aux;
    int *n_plp, dret, i, n, c, min_mapQ = 0;
    int64_t *cnt;
    const bam_pileup1_t **plp;
    int usage = 0;

    sam_global_args ga = SAM_GLOBAL_ARGS_INIT;
    static const struct option lopts[] = {
        SAM_OPT_GLOBAL_OPTIONS('-', 0, '-', '-', 0),
        { NULL, 0, NULL, 0 }
    };

    while ((c = getopt_long(argc, argv, "Q:", lopts, NULL)) >= 0) {
        switch (c) {
        case 'Q': min_mapQ = atoi(optarg); break;
        default:  if (parse_sam_global_opt(c, optarg, lopts, &ga) == 0) break;
                  /* else fall-through */
        case '?': usage = 1; break;
        }
        if (usage) break;
    }
    if (usage || optind + 2 > argc) {
        fprintf(pysam_stderr, "Usage: samtools bedcov [options] <in.bed> <in1.bam> [...]\n\n");
        fprintf(pysam_stderr, "  -Q INT       Only count bases of at least INT quality [0]\n");
        sam_global_opt_help(pysam_stderr, "-.--.");
        return 1;
    }
    memset(&str, 0, sizeof(kstring_t));
    n = argc - optind - 1;
    aux = calloc(n, sizeof(aux_t*));
    idx = calloc(n, sizeof(hts_idx_t*));
    for (i = 0; i < n; ++i) {
        aux[i] = calloc(1, sizeof(aux_t));
        aux[i]->min_mapQ = min_mapQ;
        aux[i]->fp = sam_open_format(argv[i+optind+1], "r", &ga.in);
        if (aux[i]->fp)
            idx[i] = sam_index_load(aux[i]->fp, argv[i+optind+1]);
        if (aux[i]->fp == 0 || idx[i] == 0) {
            fprintf(pysam_stderr, "ERROR: fail to open index BAM file '%s'\n", argv[i+optind+1]);
            return 2;
        }
        // TODO bgzf_set_cache_size(aux[i]->fp, 20);
        aux[i]->header = sam_hdr_read(aux[i]->fp);
        if (aux[i]->header == NULL) {
            fprintf(pysam_stderr, "ERROR: failed to read header for '%s'\n",
                    argv[i+optind+1]);
            return 2;
        }
    }
    cnt = calloc(n, 8);

    fp = gzopen(argv[optind], "rb");
    ks = ks_init(fp);
    n_plp = calloc(n, sizeof(int));
    plp = calloc(n, sizeof(bam_pileup1_t*));
    while (ks_getuntil(ks, KS_SEP_LINE, &str, &dret) >= 0) {
        char *p, *q;
        int tid, beg, end, pos;
        bam_mplp_t mplp;

        for (p = q = str.s; *p && *p != '\t'; ++p);
        if (*p != '\t') goto bed_error;
        *p = 0; tid = bam_name2id(aux[0]->header, q); *p = '\t';
        if (tid < 0) goto bed_error;
        for (q = p = p + 1; isdigit(*p); ++p);
        if (*p != '\t') goto bed_error;
        *p = 0; beg = atoi(q); *p = '\t';
        for (q = p = p + 1; isdigit(*p); ++p);
        if (*p == '\t' || *p == 0) {
            int c = *p;
            *p = 0; end = atoi(q); *p = c;
        } else goto bed_error;

        for (i = 0; i < n; ++i) {
            if (aux[i]->iter) hts_itr_destroy(aux[i]->iter);
            aux[i]->iter = sam_itr_queryi(idx[i], tid, beg, end);
        }
        mplp = bam_mplp_init(n, read_bam, (void**)aux);
        bam_mplp_set_maxcnt(mplp, 64000);
        memset(cnt, 0, 8 * n);
        while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0)
            if (pos >= beg && pos < end)
                for (i = 0; i < n; ++i) cnt[i] += n_plp[i];
        for (i = 0; i < n; ++i) {
            kputc('\t', &str);
            kputl(cnt[i], &str);
        }
        fputs(str.s, pysam_stdout) & fputc('\n', pysam_stdout);
        bam_mplp_destroy(mplp);
        continue;

bed_error:
        fprintf(pysam_stderr, "Errors in BED line '%s'\n", str.s);
    }
    free(n_plp); free(plp);
    ks_destroy(ks);
    gzclose(fp);

    free(cnt);
    for (i = 0; i < n; ++i) {
        if (aux[i]->iter) hts_itr_destroy(aux[i]->iter);
        hts_idx_destroy(idx[i]);
        bam_hdr_destroy(aux[i]->header);
        sam_close(aux[i]->fp);
        free(aux[i]);
    }
    free(aux); free(idx);
    free(str.s);
    sam_global_args_free(&ga);
    return 0;
}
Esempio n. 5
0
int main_depth(int argc, char *argv[])
{
    int i, n, tid, reg_tid, beg, end, pos, *n_plp, baseQ = 0, mapQ = 0, min_len = 0;
    int all = 0, status = EXIT_SUCCESS, nfiles, max_depth = -1;
    const bam_pileup1_t **plp;
    char *reg = 0; // specified region
    void *bed = 0; // BED data structure
    char *file_list = NULL, **fn = NULL;
    bam_hdr_t *h = NULL; // BAM header of the 1st input
    aux_t **data;
    bam_mplp_t mplp;
    int last_pos = -1, last_tid = -1, ret;

    sam_global_args ga = SAM_GLOBAL_ARGS_INIT;
    static const struct option lopts[] = {
        SAM_OPT_GLOBAL_OPTIONS('-', 0, '-', '-', 0),
        { NULL, 0, NULL, 0 }
    };

    // parse the command line
    while ((n = getopt_long(argc, argv, "r:b:q:Q:l:f:am:d:", lopts, NULL)) >= 0) {
        switch (n) {
        case 'l':
            min_len = atoi(optarg);
            break; // minimum query length
        case 'r':
            reg = strdup(optarg);
            break;   // parsing a region requires a BAM header
        case 'b':
            bed = bed_read(optarg); // BED or position list file can be parsed now
            if (!bed) {
                print_error_errno("depth", "Could not read file \"%s\"", optarg);
                return 1;
            }
            break;
        case 'q':
            baseQ = atoi(optarg);
            break;   // base quality threshold
        case 'Q':
            mapQ = atoi(optarg);
            break;    // mapping quality threshold
        case 'f':
            file_list = optarg;
            break;
        case 'a':
            all++;
            break;
        case 'd':
        case 'm':
            max_depth = atoi(optarg);
            break; // maximum coverage depth
        default:
            if (parse_sam_global_opt(n, optarg, lopts, &ga) == 0) break;
        /* else fall-through */
        case '?':
            return usage();
        }
    }
    if (optind == argc && !file_list)
        return usage();

    // initialize the auxiliary data structures
    if (file_list)
    {
        if ( read_file_list(file_list,&nfiles,&fn) ) return 1;
        n = nfiles;
        argv = fn;
        optind = 0;
    }
    else
        n = argc - optind; // the number of BAMs on the command line
    data = calloc(n, sizeof(aux_t*)); // data[i] for the i-th input
    reg_tid = 0;
    beg = 0;
    end = INT_MAX;  // set the default region
    for (i = 0; i < n; ++i) {
        int rf;
        data[i] = calloc(1, sizeof(aux_t));
        data[i]->fp = sam_open_format(argv[optind+i], "r", &ga.in); // open BAM
        if (data[i]->fp == NULL) {
            print_error_errno("depth", "Could not open \"%s\"", argv[optind+i]);
            status = EXIT_FAILURE;
            goto depth_end;
        }
        rf = SAM_FLAG | SAM_RNAME | SAM_POS | SAM_MAPQ | SAM_CIGAR | SAM_SEQ;
        if (baseQ) rf |= SAM_QUAL;
        if (hts_set_opt(data[i]->fp, CRAM_OPT_REQUIRED_FIELDS, rf)) {
            fprintf(stderr, "Failed to set CRAM_OPT_REQUIRED_FIELDS value\n");
            return 1;
        }
        if (hts_set_opt(data[i]->fp, CRAM_OPT_DECODE_MD, 0)) {
            fprintf(stderr, "Failed to set CRAM_OPT_DECODE_MD value\n");
            return 1;
        }
        data[i]->min_mapQ = mapQ;                    // set the mapQ filter
        data[i]->min_len  = min_len;                 // set the qlen filter
        data[i]->hdr = sam_hdr_read(data[i]->fp);    // read the BAM header
        if (data[i]->hdr == NULL) {
            fprintf(stderr, "Couldn't read header for \"%s\"\n",
                    argv[optind+i]);
            status = EXIT_FAILURE;
            goto depth_end;
        }
        if (reg) { // if a region is specified
            hts_idx_t *idx = sam_index_load(data[i]->fp, argv[optind+i]);  // load the index
            if (idx == NULL) {
                print_error("depth", "can't load index for \"%s\"", argv[optind+i]);
                status = EXIT_FAILURE;
                goto depth_end;
            }
            data[i]->iter = sam_itr_querys(idx, data[i]->hdr, reg); // set the iterator
            hts_idx_destroy(idx); // the index is not needed any more; free the memory
            if (data[i]->iter == NULL) {
                print_error("depth", "can't parse region \"%s\"", reg);
                status = EXIT_FAILURE;
                goto depth_end;
            }
        }
    }

    h = data[0]->hdr; // easy access to the header of the 1st BAM
    if (reg) {
        beg = data[0]->iter->beg; // and to the parsed region coordinates
        end = data[0]->iter->end;
        reg_tid = data[0]->iter->tid;
    }

    // the core multi-pileup loop
    mplp = bam_mplp_init(n, read_bam, (void**)data); // initialization
    if (0 < max_depth)
        bam_mplp_set_maxcnt(mplp,max_depth);  // set maximum coverage depth
    n_plp = calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM
    plp = calloc(n, sizeof(bam_pileup1_t*)); // plp[i] points to the array of covering reads (internal in mplp)
    while ((ret=bam_mplp_auto(mplp, &tid, &pos, n_plp, plp)) > 0) { // come to the next covered position
        if (pos < beg || pos >= end) continue; // out of range; skip
        if (tid >= h->n_targets) continue;     // diff number of @SQ lines per file?
        if (all) {
            while (tid > last_tid) {
                if (last_tid >= 0 && !reg) {
                    // Deal with remainder or entirety of last tid.
                    while (++last_pos < h->target_len[last_tid]) {
                        // Horribly inefficient, but the bed API is an obfuscated black box.
                        if (bed && bed_overlap(bed, h->target_name[last_tid], last_pos, last_pos + 1) == 0)
                            continue;
                        fputs(h->target_name[last_tid], stdout);
                        printf("\t%d", last_pos+1);
                        for (i = 0; i < n; i++)
                            putchar('\t'), putchar('0');
                        putchar('\n');
                    }
                }
                last_tid++;
                last_pos = -1;
                if (all < 2)
                    break;
            }

            // Deal with missing portion of current tid
            while (++last_pos < pos) {
                if (last_pos < beg) continue; // out of range; skip
                if (bed && bed_overlap(bed, h->target_name[tid], last_pos, last_pos + 1) == 0)
                    continue;
                fputs(h->target_name[tid], stdout);
                printf("\t%d", last_pos+1);
                for (i = 0; i < n; i++)
                    putchar('\t'), putchar('0');
                putchar('\n');
            }

            last_tid = tid;
            last_pos = pos;
        }
        if (bed && bed_overlap(bed, h->target_name[tid], pos, pos + 1) == 0) continue;
        fputs(h->target_name[tid], stdout);
        printf("\t%d", pos+1); // a customized printf() would be faster
        for (i = 0; i < n; ++i) { // base level filters have to go here
            int j, m = 0;
            for (j = 0; j < n_plp[i]; ++j) {
                const bam_pileup1_t *p = plp[i] + j; // DON'T modfity plp[][] unless you really know
                if (p->is_del || p->is_refskip) ++m; // having dels or refskips at tid:pos
                else if (bam_get_qual(p->b)[p->qpos] < baseQ) ++m; // low base quality
            }
            printf("\t%d", n_plp[i] - m); // this the depth to output
        }
        putchar('\n');
    }
    if (ret < 0) status = EXIT_FAILURE;
    free(n_plp);
    free(plp);
    bam_mplp_destroy(mplp);

    if (all) {
        // Handle terminating region
        if (last_tid < 0 && reg && all > 1) {
            last_tid = reg_tid;
            last_pos = beg-1;
        }
        while (last_tid >= 0 && last_tid < h->n_targets) {
            while (++last_pos < h->target_len[last_tid]) {
                if (last_pos >= end) break;
                if (bed && bed_overlap(bed, h->target_name[last_tid], last_pos, last_pos + 1) == 0)
                    continue;
                fputs(h->target_name[last_tid], stdout);
                printf("\t%d", last_pos+1);
                for (i = 0; i < n; i++)
                    putchar('\t'), putchar('0');
                putchar('\n');
            }
            last_tid++;
            last_pos = -1;
            if (all < 2 || reg)
                break;
        }
    }

depth_end:
    for (i = 0; i < n && data[i]; ++i) {
        bam_hdr_destroy(data[i]->hdr);
        if (data[i]->fp) sam_close(data[i]->fp);
        hts_itr_destroy(data[i]->iter);
        free(data[i]);
    }
    free(data);
    free(reg);
    if (bed) bed_destroy(bed);
    if ( file_list )
    {
        for (i=0; i<n; i++) free(fn[i]);
        free(fn);
    }
    sam_global_args_free(&ga);
    return status;
}
Esempio n. 6
0
int main_depth(int argc, char *argv[])
#endif
{
	int i, n, tid, beg, end, pos, *n_plp, baseQ = 0, mapQ = 0, min_len = 0, nfiles;
	const bam_pileup1_t **plp;
	char *reg = 0; // specified region
	void *bed = 0; // BED data structure
    char *file_list = NULL, **fn = NULL;
	bam_header_t *h = 0; // BAM header of the 1st input
	aux_t **data;
	bam_mplp_t mplp;

	// parse the command line
	while ((n = getopt(argc, argv, "r:b:q:Q:l:f:")) >= 0) {
		switch (n) {
			case 'l': min_len = atoi(optarg); break; // minimum query length
			case 'r': reg = strdup(optarg); break;   // parsing a region requires a BAM header
			case 'b': bed = bed_read(optarg); break; // BED or position list file can be parsed now
			case 'q': baseQ = atoi(optarg); break;   // base quality threshold
			case 'Q': mapQ = atoi(optarg); break;    // mapping quality threshold
			case 'f': file_list = optarg; break;
		}
	}
	if (optind == argc && !file_list) {
        fprintf(stderr, "\n");
        fprintf(stderr, "Usage: samtools depth [options] in1.bam [in2.bam [...]]\n");
        fprintf(stderr, "Options:\n");
        fprintf(stderr, "   -b <bed>            list of positions or regions\n");
        fprintf(stderr, "   -f <list>           list of input BAM filenames, one per line [null]\n");
        fprintf(stderr, "   -l <int>            minQLen\n");
        fprintf(stderr, "   -q <int>            base quality threshold\n");
        fprintf(stderr, "   -Q <int>            mapping quality threshold\n");
        fprintf(stderr, "   -r <chr:from-to>    region\n");
        fprintf(stderr, "\n");
		return 1;
	}

	// initialize the auxiliary data structures
    if (file_list) 
    {
        if ( read_file_list(file_list,&nfiles,&fn) ) return 1;
        n = nfiles;
        argv = fn;
        optind = 0;
    }
    else
        n = argc - optind; // the number of BAMs on the command line
	data = calloc(n, sizeof(void*)); // data[i] for the i-th input
	beg = 0; end = 1<<30; tid = -1;  // set the default region
	for (i = 0; i < n; ++i) {
		bam_header_t *htmp;
		data[i] = calloc(1, sizeof(aux_t));
		data[i]->fp = bam_open(argv[optind+i], "r"); // open BAM
		data[i]->min_mapQ = mapQ;                    // set the mapQ filter
		data[i]->min_len  = min_len;                 // set the qlen filter
		htmp = bam_header_read(data[i]->fp);         // read the BAM header
		if (i == 0) {
			h = htmp; // keep the header of the 1st BAM
			if (reg) bam_parse_region(h, reg, &tid, &beg, &end); // also parse the region
		} else bam_header_destroy(htmp); // if not the 1st BAM, trash the header
		if (tid >= 0) { // if a region is specified and parsed successfully
			bam_index_t *idx = bam_index_load(argv[optind+i]);  // load the index
			data[i]->iter = bam_iter_query(idx, tid, beg, end); // set the iterator
			bam_index_destroy(idx); // the index is not needed any more; phase out of the memory
		}
	}

	// the core multi-pileup loop
	mplp = bam_mplp_init(n, read_bam, (void**)data); // initialization
	bam_mplp_set_maxcnt(mplp,2147483647); // set max_depth to int max
	n_plp = calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM
	plp = calloc(n, sizeof(void*)); // plp[i] points to the array of covering reads (internal in mplp)
	while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0) { // come to the next covered position
		if (pos < beg || pos >= end) continue; // out of range; skip
		if (bed && bed_overlap(bed, h->target_name[tid], pos, pos + 1) == 0) continue; // not in BED; skip
		fputs(h->target_name[tid], stdout); printf("\t%d", pos+1); // a customized printf() would be faster
		for (i = 0; i < n; ++i) { // base level filters have to go here
			int j, m = 0;
			for (j = 0; j < n_plp[i]; ++j) {
				const bam_pileup1_t *p = plp[i] + j; // DON'T modfity plp[][] unless you really know
				if (p->is_del || p->is_refskip) ++m; // having dels or refskips at tid:pos
				else if (bam1_qual(p->b)[p->qpos] < baseQ) ++m; // low base quality
			}
			printf("\t%d", n_plp[i] - m); // this the depth to output
		}
		putchar('\n');
	}
	free(n_plp); free(plp);
	bam_mplp_destroy(mplp);

	bam_header_destroy(h);
	for (i = 0; i < n; ++i) {
		bam_close(data[i]->fp);
		if (data[i]->iter) bam_iter_destroy(data[i]->iter);
		free(data[i]);
	}
	free(data); free(reg);
	if (bed) bed_destroy(bed);
    if ( file_list )
    {
        for (i=0; i<n; i++) free(fn[i]);
        free(fn);
    }
	return 0;
}
Esempio n. 7
0
static int mpileup(mplp_conf_t *conf)
{
    if (conf->nfiles == 0) {
        fprintf(stderr,"[%s] no input file/data given\n", __func__);
        exit(EXIT_FAILURE);
    }

    mplp_ref_t mp_ref = MPLP_REF_INIT;
    conf->gplp = (mplp_pileup_t *) calloc(1,sizeof(mplp_pileup_t));
    conf->mplp_data = (mplp_aux_t**) calloc(conf->nfiles, sizeof(mplp_aux_t*));
    conf->plp = (const bam_pileup1_t**) calloc(conf->nfiles, sizeof(bam_pileup1_t*));
    conf->n_plp = (int*) calloc(conf->nfiles, sizeof(int));

    // Allow to run mpileup on multiple regions in one go. This comes at cost: the bai index
    // must be kept in the memory for the whole time which can be a problem with many bams.
    // Therefore if none or only one region is requested, we initialize the bam iterator as
    // before and free the index. Only when multiple regions are queried, we keep the index.
    int nregs = 0;
    if ( conf->reg_fname )
    {
        if ( conf->reg_is_file )
        {
            conf->reg = regidx_init(conf->reg_fname,NULL,NULL,0,NULL);
            if ( !conf->reg ) {
                fprintf(stderr,"Could not parse the regions: %s\n", conf->reg_fname);
                exit(EXIT_FAILURE);
            }
        }
        else
        {
            conf->reg = regidx_init(NULL,regidx_parse_reg,NULL,sizeof(char*),NULL);
            if ( regidx_insert_list(conf->reg,conf->reg_fname,',') !=0 ) {
                fprintf(stderr,"Could not parse the regions: %s\n", conf->reg_fname);
                exit(EXIT_FAILURE);
            }
        }
        nregs = regidx_nregs(conf->reg);
        conf->reg_itr = regitr_init(conf->reg);
        regitr_loop(conf->reg_itr);   // region iterator now positioned at the first region
    }

    // read the header of each file in the list and initialize data
    // beware: mpileup has always assumed that tid's are consistent in the headers, add sanity check at least!
    bam_hdr_t *hdr = NULL;      // header of first file in input list
    int i;
    for (i = 0; i < conf->nfiles; ++i) {
        bam_hdr_t *h_tmp;
        conf->mplp_data[i] = (mplp_aux_t*) calloc(1, sizeof(mplp_aux_t));
        conf->mplp_data[i]->fp = sam_open(conf->files[i], "rb");
        if ( !conf->mplp_data[i]->fp )
        {
            fprintf(stderr, "[%s] failed to open %s: %s\n", __func__, conf->files[i], strerror(errno));
            exit(EXIT_FAILURE);
        }
        if (hts_set_opt(conf->mplp_data[i]->fp, CRAM_OPT_DECODE_MD, 0)) {
            fprintf(stderr, "Failed to set CRAM_OPT_DECODE_MD value\n");
            exit(EXIT_FAILURE);
        }
        if (conf->fai_fname && hts_set_fai_filename(conf->mplp_data[i]->fp, conf->fai_fname) != 0) {
            fprintf(stderr, "[%s] failed to process %s: %s\n",
                    __func__, conf->fai_fname, strerror(errno));
            exit(EXIT_FAILURE);
        }
        conf->mplp_data[i]->conf = conf;
        conf->mplp_data[i]->ref = &mp_ref;
        h_tmp = sam_hdr_read(conf->mplp_data[i]->fp);
        if ( !h_tmp ) {
            fprintf(stderr,"[%s] fail to read the header of %s\n", __func__, conf->files[i]);
            exit(EXIT_FAILURE);
        }
        conf->mplp_data[i]->h = i ? hdr : h_tmp; // for j==0, "h" has not been set yet
        conf->mplp_data[i]->bam_id = bam_smpl_add_bam(conf->bsmpl,h_tmp->text,conf->files[i]);
        if ( conf->mplp_data[i]->bam_id<0 )
        {
            // no usable readgroups in this bam, it can be skipped
            sam_close(conf->mplp_data[i]->fp);
            free(conf->mplp_data[i]);
            bam_hdr_destroy(h_tmp);
            free(conf->files[i]);
            if ( i+1<conf->nfiles ) memmove(&conf->files[i],&conf->files[i+1],sizeof(*conf->files)*(conf->nfiles-i-1));
            conf->nfiles--;
            i--;
            continue;
        }
        if (conf->reg) {
            hts_idx_t *idx = sam_index_load(conf->mplp_data[i]->fp, conf->files[i]);
            if (idx == NULL) {
                fprintf(stderr, "[%s] fail to load index for %s\n", __func__, conf->files[i]);
                exit(EXIT_FAILURE);
            }
            conf->buf.l = 0;
            ksprintf(&conf->buf,"%s:%u-%u",conf->reg_itr->seq,conf->reg_itr->beg+1,conf->reg_itr->end+1);
            conf->mplp_data[i]->iter = sam_itr_querys(idx, conf->mplp_data[i]->h, conf->buf.s);
            if ( !conf->mplp_data[i]->iter ) 
            {
                conf->mplp_data[i]->iter = sam_itr_querys(idx, conf->mplp_data[i]->h, conf->reg_itr->seq);
                if ( conf->mplp_data[i]->iter ) {
                    fprintf(stderr,"[E::%s] fail to parse region '%s'\n", __func__, conf->buf.s);
                    exit(EXIT_FAILURE);
                }
                fprintf(stderr,"[E::%s] the sequence \"%s\" not found: %s\n",__func__,conf->reg_itr->seq,conf->files[i]);
                exit(EXIT_FAILURE);
            }
            if ( nregs==1 ) // no need to keep the index in memory
               hts_idx_destroy(idx);
            else
                conf->mplp_data[i]->idx = idx;
        }

        if ( !hdr ) hdr = h_tmp; /* save the header of first file in list */
        else {
            // FIXME: check consistency between h and h_tmp
            bam_hdr_destroy(h_tmp);

            // we store only the first file's header; it's (alleged to be)
            // compatible with the i-th file's target_name lookup needs
            conf->mplp_data[i]->h = hdr;
        }
    }
    // allocate data storage proportionate to number of samples being studied sm->n
    bam_smpl_get_samples(conf->bsmpl, &conf->gplp->n);
    conf->gplp->n_plp = (int*) calloc(conf->gplp->n, sizeof(int));
    conf->gplp->m_plp = (int*) calloc(conf->gplp->n, sizeof(int));
    conf->gplp->plp = (bam_pileup1_t**) calloc(conf->gplp->n, sizeof(bam_pileup1_t*));  

    fprintf(stderr, "[%s] %d samples in %d input files\n", __func__, conf->gplp->n, conf->nfiles);
    // write the VCF header
    conf->bcf_fp = hts_open(conf->output_fname?conf->output_fname:"-", hts_bcf_wmode(conf->output_type));
    if (conf->bcf_fp == NULL) {
        fprintf(stderr, "[%s] failed to write to %s: %s\n", __func__, conf->output_fname? conf->output_fname : "standard output", strerror(errno));
        exit(EXIT_FAILURE);
    }
    if ( conf->n_threads ) hts_set_threads(conf->bcf_fp, conf->n_threads);

    // BCF header creation
    conf->bcf_hdr = bcf_hdr_init("w");
    conf->buf.l = 0;

    if (conf->record_cmd_line)
    {
        ksprintf(&conf->buf, "##bcftoolsVersion=%s+htslib-%s\n",bcftools_version(),hts_version());
        bcf_hdr_append(conf->bcf_hdr, conf->buf.s);

        conf->buf.l = 0;
        ksprintf(&conf->buf, "##bcftoolsCommand=mpileup");
        for (i=1; i<conf->argc; i++) ksprintf(&conf->buf, " %s", conf->argv[i]);
        kputc('\n', &conf->buf);
        bcf_hdr_append(conf->bcf_hdr, conf->buf.s);
    }

    if (conf->fai_fname)
    {
        conf->buf.l = 0;
        ksprintf(&conf->buf, "##reference=file://%s\n", conf->fai_fname);
        bcf_hdr_append(conf->bcf_hdr, conf->buf.s);
    }

    // Translate BAM @SQ tags to BCF ##contig tags
    // todo: use/write new BAM header manipulation routines, fill also UR, M5
    for (i=0; i<hdr->n_targets; i++)
    {
        conf->buf.l = 0;
        ksprintf(&conf->buf, "##contig=<ID=%s,length=%d>", hdr->target_name[i], hdr->target_len[i]);
        bcf_hdr_append(conf->bcf_hdr, conf->buf.s);
    }
    conf->buf.l = 0;

    bcf_hdr_append(conf->bcf_hdr,"##ALT=<ID=*,Description=\"Represents allele(s) other than observed.\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=INDEL,Number=0,Type=Flag,Description=\"Indicates that the variant is an INDEL.\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=IDV,Number=1,Type=Integer,Description=\"Maximum number of reads supporting an indel\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=IMF,Number=1,Type=Float,Description=\"Maximum fraction of reads supporting an indel\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=DP,Number=1,Type=Integer,Description=\"Raw read depth\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=VDB,Number=1,Type=Float,Description=\"Variant Distance Bias for filtering splice-site artefacts in RNA-seq data (bigger is better)\",Version=\"3\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=RPB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Read Position Bias (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=MQB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality Bias (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=BQB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Base Quality Bias (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=MQSB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality vs Strand Bias (bigger is better)\">");
#if CDF_MWU_TESTS
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=RPB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Read Position Bias [CDF] (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=MQB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality Bias [CDF] (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=BQB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Base Quality Bias [CDF] (bigger is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=MQSB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality vs Strand Bias [CDF] (bigger is better)\">");
#endif
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=SGB,Number=1,Type=Float,Description=\"Segregation based metric.\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=MQ0F,Number=1,Type=Float,Description=\"Fraction of MQ0 reads (smaller is better)\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=I16,Number=16,Type=Float,Description=\"Auxiliary tag used for calling, see description of bcf_callret1_t in bam2bcf.h\">");
    bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=QS,Number=R,Type=Float,Description=\"Auxiliary tag used for calling\">");
    bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=PL,Number=G,Type=Integer,Description=\"List of Phred-scaled genotype likelihoods\">");
    if ( conf->fmt_flag&B2B_FMT_DP )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=DP,Number=1,Type=Integer,Description=\"Number of high-quality bases\">");
    if ( conf->fmt_flag&B2B_FMT_DV )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=DV,Number=1,Type=Integer,Description=\"Number of high-quality non-reference bases\">");
    if ( conf->fmt_flag&B2B_FMT_DPR )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=DPR,Number=R,Type=Integer,Description=\"Number of high-quality bases observed for each allele\">");
    if ( conf->fmt_flag&B2B_INFO_DPR )
        bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=DPR,Number=R,Type=Integer,Description=\"Number of high-quality bases observed for each allele\">");
    if ( conf->fmt_flag&B2B_FMT_DP4 )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=DP4,Number=4,Type=Integer,Description=\"Number of high-quality ref-fwd, ref-reverse, alt-fwd and alt-reverse bases\">");
    if ( conf->fmt_flag&B2B_FMT_SP )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=SP,Number=1,Type=Integer,Description=\"Phred-scaled strand bias P-value\">");
    if ( conf->fmt_flag&B2B_FMT_AD )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=AD,Number=R,Type=Integer,Description=\"Allelic depths\">");
    if ( conf->fmt_flag&B2B_FMT_ADF )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=ADF,Number=R,Type=Integer,Description=\"Allelic depths on the forward strand\">");
    if ( conf->fmt_flag&B2B_FMT_ADR )
        bcf_hdr_append(conf->bcf_hdr,"##FORMAT=<ID=ADR,Number=R,Type=Integer,Description=\"Allelic depths on the reverse strand\">");
    if ( conf->fmt_flag&B2B_INFO_AD )
        bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=AD,Number=R,Type=Integer,Description=\"Total allelic depths\">");
    if ( conf->fmt_flag&B2B_INFO_ADF )
        bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=ADF,Number=R,Type=Integer,Description=\"Total allelic depths on the forward strand\">");
    if ( conf->fmt_flag&B2B_INFO_ADR )
        bcf_hdr_append(conf->bcf_hdr,"##INFO=<ID=ADR,Number=R,Type=Integer,Description=\"Total allelic depths on the reverse strand\">");
    if ( conf->gvcf )
        gvcf_update_header(conf->gvcf, conf->bcf_hdr);

    int nsmpl;
    const char **smpl = bam_smpl_get_samples(conf->bsmpl, &nsmpl);
    for (i=0; i<nsmpl; i++)
        bcf_hdr_add_sample(conf->bcf_hdr, smpl[i]);
    bcf_hdr_write(conf->bcf_fp, conf->bcf_hdr);

    conf->bca = bcf_call_init(-1., conf->min_baseQ);
    conf->bcr = (bcf_callret1_t*) calloc(nsmpl, sizeof(bcf_callret1_t));
    conf->bca->openQ = conf->openQ, conf->bca->extQ = conf->extQ, conf->bca->tandemQ = conf->tandemQ;
    conf->bca->min_frac = conf->min_frac;
    conf->bca->min_support = conf->min_support;
    conf->bca->per_sample_flt = conf->flag & MPLP_PER_SAMPLE;

    conf->bc.bcf_hdr = conf->bcf_hdr;
    conf->bc.n  = nsmpl;
    conf->bc.PL = (int32_t*) malloc(15 * nsmpl * sizeof(*conf->bc.PL));
    if (conf->fmt_flag)
    {
        assert( sizeof(float)==sizeof(int32_t) );
        conf->bc.DP4 = (int32_t*) malloc(nsmpl * sizeof(int32_t) * 4);
        conf->bc.fmt_arr = (uint8_t*) malloc(nsmpl * sizeof(float)); // all fmt_flag fields, float and int32
        if ( conf->fmt_flag&(B2B_INFO_DPR|B2B_FMT_DPR|B2B_INFO_AD|B2B_INFO_ADF|B2B_INFO_ADR|B2B_FMT_AD|B2B_FMT_ADF|B2B_FMT_ADR) )
        {
            // first B2B_MAX_ALLELES fields for total numbers, the rest per-sample
            conf->bc.ADR = (int32_t*) malloc((nsmpl+1)*B2B_MAX_ALLELES*sizeof(int32_t));
            conf->bc.ADF = (int32_t*) malloc((nsmpl+1)*B2B_MAX_ALLELES*sizeof(int32_t));
            for (i=0; i<nsmpl; i++)
            {
                conf->bcr[i].ADR = conf->bc.ADR + (i+1)*B2B_MAX_ALLELES;
                conf->bcr[i].ADF = conf->bc.ADF + (i+1)*B2B_MAX_ALLELES;
            }
        }
    }

    // init mpileup
    conf->iter = bam_mplp_init(conf->nfiles, mplp_func, (void**)conf->mplp_data);
    if ( conf->flag & MPLP_SMART_OVERLAPS ) bam_mplp_init_overlaps(conf->iter);
    if ( (double)conf->max_depth * conf->nfiles > 1<<20)
        fprintf(stderr, "Warning: Potential memory hog, up to %.0fM reads in the pileup!\n", (double)conf->max_depth*conf->nfiles);
    if ( (double)conf->max_depth * conf->nfiles / nsmpl < 250 )
        fprintf(stderr, "Note: The maximum per-sample depth with -d %d is %.1fx\n", conf->max_depth,(double)conf->max_depth * conf->nfiles / nsmpl);
    bam_mplp_set_maxcnt(conf->iter, conf->max_depth);
    conf->max_indel_depth = conf->max_indel_depth * nsmpl;
    conf->bcf_rec = bcf_init1();
    bam_mplp_constructor(conf->iter, pileup_constructor);

    // Run mpileup for multiple regions
    if ( nregs )
    {
        int ireg = 0;
        do 
        {
            // first region is already positioned
            if ( ireg++ > 0 )
            {
                conf->buf.l = 0;
                ksprintf(&conf->buf,"%s:%u-%u",conf->reg_itr->seq,conf->reg_itr->beg,conf->reg_itr->end);

                for (i=0; i<conf->nfiles; i++) 
                {
                    hts_itr_destroy(conf->mplp_data[i]->iter);
                    conf->mplp_data[i]->iter = sam_itr_querys(conf->mplp_data[i]->idx, conf->mplp_data[i]->h, conf->buf.s);
                    if ( !conf->mplp_data[i]->iter ) 
                    {
                        conf->mplp_data[i]->iter = sam_itr_querys(conf->mplp_data[i]->idx, conf->mplp_data[i]->h, conf->reg_itr->seq);
                        if ( conf->mplp_data[i]->iter ) {
                            fprintf(stderr,"[E::%s] fail to parse region '%s'\n", __func__, conf->buf.s);
                            exit(EXIT_FAILURE);
                        }
                        fprintf(stderr,"[E::%s] the sequence \"%s\" not found: %s\n",__func__,conf->reg_itr->seq,conf->files[i]);
                        exit(EXIT_FAILURE);
                    }
                    bam_mplp_reset(conf->iter);
                }
            }
            mpileup_reg(conf,conf->reg_itr->beg,conf->reg_itr->end);
        }
        while ( regitr_loop(conf->reg_itr) );
    }
    else
        mpileup_reg(conf,0,0);

    flush_bcf_records(conf, conf->bcf_fp, conf->bcf_hdr, NULL);

    // clean up
    free(conf->bc.tmp.s);
    bcf_destroy1(conf->bcf_rec);
    if (conf->bcf_fp)
    {
        hts_close(conf->bcf_fp);
        bcf_hdr_destroy(conf->bcf_hdr);
        bcf_call_destroy(conf->bca);
        free(conf->bc.PL);
        free(conf->bc.DP4);
        free(conf->bc.ADR);
        free(conf->bc.ADF);
        free(conf->bc.fmt_arr);
        free(conf->bcr);
    }
    if ( conf->gvcf ) gvcf_destroy(conf->gvcf);
    free(conf->buf.s);
    for (i = 0; i < conf->gplp->n; ++i) free(conf->gplp->plp[i]);
    free(conf->gplp->plp); free(conf->gplp->n_plp); free(conf->gplp->m_plp); free(conf->gplp);
    bam_mplp_destroy(conf->iter);
    bam_hdr_destroy(hdr);
    for (i = 0; i < conf->nfiles; ++i) {
        if ( nregs>1 ) hts_idx_destroy(conf->mplp_data[i]->idx);
        sam_close(conf->mplp_data[i]->fp);
        if ( conf->mplp_data[i]->iter) hts_itr_destroy(conf->mplp_data[i]->iter);
        free(conf->mplp_data[i]);
    }
    if ( conf->reg_itr ) regitr_destroy(conf->reg_itr);
    free(conf->mplp_data); free(conf->plp); free(conf->n_plp);
    free(mp_ref.ref[0]);
    free(mp_ref.ref[1]);
    return 0;
}
Esempio n. 8
0
/*
 * Performs pileup
 * @param conf configuration for this pileup
 * @param n number of files specified in fn
 * @param fn filenames
 */
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, tid0 = -1, beg0 = 0, end0 = 1u<<29, ref_len, ref_tid = -1, max_depth, max_indel_depth;
    const bam_pileup1_t **plp;
    bam_mplp_t iter;
    bam_hdr_t *h = NULL; /* header of first file in input list */
    char *ref;
    void *rghash = NULL;
    FILE *pileup_fp = NULL;

    bcf_callaux_t *bca = NULL;
    bcf_callret1_t *bcr = NULL;
    bcf_call_t bc;
    htsFile *bcf_fp = NULL;
    bcf_hdr_t *bcf_hdr = NULL;

    bam_sample_t *sm = NULL;
    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(mplp_aux_t*));
    plp = calloc(n, sizeof(bam_pileup1_t*));
    n_plp = calloc(n, sizeof(int));
    sm = bam_smpl_init();

    if (n == 0) {
        fprintf(stderr,"[%s] no input file/data given\n", __func__);
        exit(1);
    }

    // read the header of each file in the list and initialize data
    for (i = 0; i < n; ++i) {
        bam_hdr_t *h_tmp;
        data[i] = calloc(1, sizeof(mplp_aux_t));
        data[i]->fp = sam_open(fn[i], "rb");
        if ( !data[i]->fp )
        {
            fprintf(stderr, "[%s] failed to open %s: %s\n", __func__, fn[i], strerror(errno));
            exit(1);
        }
        hts_set_fai_filename(data[i]->fp, conf->fai_fname);
        data[i]->conf = conf;
        h_tmp = sam_hdr_read(data[i]->fp);
        if ( !h_tmp ) {
            fprintf(stderr,"[%s] fail to read the header of %s\n", __func__, fn[i]);
            exit(1);
        }
        data[i]->h = i? h : h_tmp; // for i==0, "h" has not been set yet
        bam_smpl_add(sm, fn[i], (conf->flag&MPLP_IGNORE_RG)? 0 : h_tmp->text);
        // Collect read group IDs with PL (platform) listed in pl_list (note: fragile, strstr search)
        rghash = bcf_call_add_rg(rghash, h_tmp->text, conf->pl_list);
        if (conf->reg) {
            hts_idx_t *idx = sam_index_load(data[i]->fp, fn[i]);
            if (idx == 0) {
                fprintf(stderr, "[%s] fail to load index for %s\n", __func__, fn[i]);
                exit(1);
            }
            if ( (data[i]->iter=sam_itr_querys(idx, data[i]->h, conf->reg)) == 0) {
                fprintf(stderr, "[E::%s] fail to parse region '%s'\n", __func__, conf->reg);
                exit(1);
            }
            if (i == 0) tid0 = data[i]->iter->tid, beg0 = data[i]->iter->beg, end0 = data[i]->iter->end;
            hts_idx_destroy(idx);
        }
        if (i == 0) h = h_tmp; /* save the header of first file in list */
        else {
            // FIXME: to check consistency
            bam_hdr_destroy(h_tmp);
        }
    }
    // allocate data storage proportionate to number of samples being studied sm->n
    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(bam_pileup1_t*));

    fprintf(stderr, "[%s] %d samples in %d input files\n", __func__, sm->n, n);
    // write the VCF header
    if (conf->flag & MPLP_BCF)
    {
        const char *mode;
        if ( conf->flag & MPLP_VCF )
            mode = (conf->flag&MPLP_NO_COMP)? "wu" : "wz";   // uncompressed VCF or compressed VCF
        else
            mode = (conf->flag&MPLP_NO_COMP)? "wub" : "wb";  // uncompressed BCF or compressed BCF

        bcf_fp = bcf_open(conf->output_fname? conf->output_fname : "-", mode);
        if (bcf_fp == NULL) {
            fprintf(stderr, "[%s] failed to write to %s: %s\n", __func__, conf->output_fname? conf->output_fname : "standard output", strerror(errno));
            exit(1);
        }

        bcf_hdr = bcf_hdr_init("w");
        kstring_t str = {0,0,0};

        ksprintf(&str, "##samtoolsVersion=%s+htslib-%s\n",samtools_version(),hts_version());
        bcf_hdr_append(bcf_hdr, str.s);

        str.l = 0;
        ksprintf(&str, "##samtoolsCommand=samtools mpileup");
        for (i=1; i<conf->argc; i++) ksprintf(&str, " %s", conf->argv[i]);
        kputc('\n', &str);
        bcf_hdr_append(bcf_hdr, str.s);

        if (conf->fai_fname)
        {
            str.l = 0;
            ksprintf(&str, "##reference=file://%s\n", conf->fai_fname);
            bcf_hdr_append(bcf_hdr, str.s);
        }

        // todo: use/write new BAM header manipulation routines, fill also UR, M5
        for (i=0; i<h->n_targets; i++)
        {
            str.l = 0;
            ksprintf(&str, "##contig=<ID=%s,length=%d>", h->target_name[i], h->target_len[i]);
            bcf_hdr_append(bcf_hdr, str.s);
        }
        free(str.s);
        bcf_hdr_append(bcf_hdr,"##ALT=<ID=X,Description=\"Represents allele(s) other than observed.\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=INDEL,Number=0,Type=Flag,Description=\"Indicates that the variant is an INDEL.\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=IDV,Number=1,Type=Integer,Description=\"Maximum number of reads supporting an indel\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=IMF,Number=1,Type=Float,Description=\"Maximum fraction of reads supporting an indel\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=DP,Number=1,Type=Integer,Description=\"Raw read depth\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=VDB,Number=1,Type=Float,Description=\"Variant Distance Bias for filtering splice-site artefacts in RNA-seq data (bigger is better)\",Version=\"3\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=RPB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Read Position Bias (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=MQB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality Bias (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=BQB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Base Quality Bias (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=MQSB,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality vs Strand Bias (bigger is better)\">");
#if CDF_MWU_TESTS
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=RPB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Read Position Bias [CDF] (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=MQB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality Bias [CDF] (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=BQB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Base Quality Bias [CDF] (bigger is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=MQSB2,Number=1,Type=Float,Description=\"Mann-Whitney U test of Mapping Quality vs Strand Bias [CDF] (bigger is better)\">");
#endif
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=SGB,Number=1,Type=Float,Description=\"Segregation based metric.\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=MQ0F,Number=1,Type=Float,Description=\"Fraction of MQ0 reads (smaller is better)\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=I16,Number=16,Type=Float,Description=\"Auxiliary tag used for calling, see description of bcf_callret1_t in bam2bcf.h\">");
        bcf_hdr_append(bcf_hdr,"##INFO=<ID=QS,Number=R,Type=Float,Description=\"Auxiliary tag used for calling\">");
        bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=PL,Number=G,Type=Integer,Description=\"List of Phred-scaled genotype likelihoods\">");
        if ( conf->fmt_flag&B2B_FMT_DP )
            bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=DP,Number=1,Type=Integer,Description=\"Number of high-quality bases\">");
        if ( conf->fmt_flag&B2B_FMT_DV )
            bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=DV,Number=1,Type=Integer,Description=\"Number of high-quality non-reference bases\">");
        if ( conf->fmt_flag&B2B_FMT_DPR )
            bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=DPR,Number=R,Type=Integer,Description=\"Number of high-quality bases observed for each allele\">");
        if ( conf->fmt_flag&B2B_INFO_DPR )
            bcf_hdr_append(bcf_hdr,"##INFO=<ID=DPR,Number=R,Type=Integer,Description=\"Number of high-quality bases observed for each allele\">");
        if ( conf->fmt_flag&B2B_FMT_DP4 )
            bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=DP4,Number=4,Type=Integer,Description=\"Number of high-quality ref-fwd, ref-reverse, alt-fwd and alt-reverse bases\">");
        if ( conf->fmt_flag&B2B_FMT_SP )
            bcf_hdr_append(bcf_hdr,"##FORMAT=<ID=SP,Number=1,Type=Integer,Description=\"Phred-scaled strand bias P-value\">");

        for (i=0; i<sm->n; i++)
            bcf_hdr_add_sample(bcf_hdr, sm->smpl[i]);
        bcf_hdr_add_sample(bcf_hdr, NULL);
        bcf_hdr_write(bcf_fp, bcf_hdr);

        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;
        bca->per_sample_flt = conf->flag & MPLP_PER_SAMPLE;

        bc.bcf_hdr = bcf_hdr;
        bc.n = sm->n;
        bc.PL = malloc(15 * sm->n * sizeof(*bc.PL));
        if (conf->fmt_flag)
        {
            assert( sizeof(float)==sizeof(int32_t) );
            bc.DP4 = malloc(sm->n * sizeof(int32_t) * 4);
            bc.fmt_arr = malloc(sm->n * sizeof(float)); // all fmt_flag fields
            if ( conf->fmt_flag&(B2B_INFO_DPR|B2B_FMT_DPR) )
            {
                // first B2B_MAX_ALLELES fields for total numbers, the rest per-sample
                bc.DPR = malloc((sm->n+1)*B2B_MAX_ALLELES*sizeof(int32_t));
                for (i=0; i<sm->n; i++)
                    bcr[i].DPR = bc.DPR + (i+1)*B2B_MAX_ALLELES;
            }
        }
    }
    else {
        pileup_fp = conf->output_fname? fopen(conf->output_fname, "w") : stdout;

        if (pileup_fp == NULL) {
            fprintf(stderr, "[%s] failed to write to %s: %s\n", __func__, conf->output_fname, strerror(errno));
            exit(1);
        }
    }

    if (tid0 >= 0 && conf->fai) { // region is set
        ref = faidx_fetch_seq(conf->fai, h->target_name[tid0], 0, 0x7fffffff, &ref_len);
        ref_tid = tid0;
        for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid0;
    } else ref_tid = -1, ref = 0;

    // begin pileup
    iter = bam_mplp_init(n, mplp_func, (void**)data);
    if ( conf->flag & MPLP_SMART_OVERLAPS ) bam_mplp_init_overlaps(iter);
    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);
    bcf1_t *bcf_rec = bcf_init1();
    int ret;
    while ( (ret=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); ref = 0;
            if (conf->fai) ref = faidx_fetch_seq(conf->fai, h->target_name[tid], 0, 0x7fffffff, &ref_len);
            for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid;
            ref_tid = tid;
        }
        if (conf->flag & MPLP_BCF) {
            int total_depth, _ref0, ref16;
            for (i = total_depth = 0; i < n; ++i) total_depth += n_plp[i];
            group_smpl(&gplp, sm, &buf, n, fn, n_plp, plp, conf->flag & MPLP_IGNORE_RG);
            _ref0 = (ref && pos < ref_len)? ref[pos] : 'N';
            ref16 = seq_nt16_table[_ref0];
            bcf_callaux_clean(bca, &bc);
            for (i = 0; i < gplp.n; ++i)
                bcf_call_glfgen(gplp.n_plp[i], gplp.plp[i], ref16, bca, bcr + i);
            bc.tid = tid; bc.pos = pos;
            bcf_call_combine(gplp.n, bcr, bca, ref16, &bc);
            bcf_clear1(bcf_rec);
            bcf_call2bcf(&bc, bcf_rec, bcr, conf->fmt_flag, 0, 0);
            bcf_write1(bcf_fp, bcf_hdr, bcf_rec);
            // call indels; todo: subsampling with total_depth>max_indel_depth instead of ignoring?
            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)
            {
                bcf_callaux_clean(bca, &bc);
                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, bca, -1, &bc) >= 0) {
                    bcf_clear1(bcf_rec);
                    bcf_call2bcf(&bc, bcf_rec, bcr, conf->fmt_flag, bca, ref);
                    bcf_write1(bcf_fp, bcf_hdr, bcf_rec);
                }
            }
        } else {
            fprintf(pileup_fp, "%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, cnt;
                for (j = cnt = 0; j < n_plp[i]; ++j) {
                    const bam_pileup1_t *p = plp[i] + j;
                    if (bam_get_qual(p->b)[p->qpos] >= conf->min_baseQ) ++cnt;
                }
                fprintf(pileup_fp, "\t%d\t", cnt);
                if (n_plp[i] == 0) {
                    fputs("*\t*", pileup_fp);
                    if (conf->flag & MPLP_PRINT_MAPQ) fputs("\t*", pileup_fp);
                    if (conf->flag & MPLP_PRINT_POS) fputs("\t*", pileup_fp);
                } else {
                    for (j = 0; j < n_plp[i]; ++j) {
                        const bam_pileup1_t *p = plp[i] + j;
                        if (bam_get_qual(p->b)[p->qpos] >= conf->min_baseQ)
                            pileup_seq(pileup_fp, plp[i] + j, pos, ref_len, ref);
                    }
                    putc('\t', pileup_fp);
                    for (j = 0; j < n_plp[i]; ++j) {
                        const bam_pileup1_t *p = plp[i] + j;
                        int c = bam_get_qual(p->b)[p->qpos];
                        if (c >= conf->min_baseQ) {
                            c = c + 33 < 126? c + 33 : 126;
                            putc(c, pileup_fp);
                        }
                    }
                    if (conf->flag & MPLP_PRINT_MAPQ) {
                        putc('\t', pileup_fp);
                        for (j = 0; j < n_plp[i]; ++j) {
                            const bam_pileup1_t *p = plp[i] + j;
                            int c = bam_get_qual(p->b)[p->qpos];
                            if ( c < conf->min_baseQ ) continue;
                            c = plp[i][j].b->core.qual + 33;
                            if (c > 126) c = 126;
                            putc(c, pileup_fp);
                        }
                    }
                    if (conf->flag & MPLP_PRINT_POS) {
                        putc('\t', pileup_fp);
                        for (j = 0; j < n_plp[i]; ++j) {
                            if (j > 0) putc(',', pileup_fp);
                            fprintf(pileup_fp, "%d", plp[i][j].qpos + 1); // FIXME: printf() is very slow...
                        }
                    }
                }
            }
            putc('\n', pileup_fp);
        }
    }

    // clean up
    free(bc.tmp.s);
    bcf_destroy1(bcf_rec);
    if (bcf_fp)
    {
        hts_close(bcf_fp);
        bcf_hdr_destroy(bcf_hdr);
        bcf_call_destroy(bca);
        free(bc.PL);
        free(bc.DP4);
        free(bc.DPR);
        free(bc.fmt_arr);
        free(bcr);
    }
    if (pileup_fp && conf->output_fname) fclose(pileup_fp);
    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);
    bam_mplp_destroy(iter);
    bam_hdr_destroy(h);
    for (i = 0; i < n; ++i) {
        sam_close(data[i]->fp);
        if (data[i]->iter) hts_itr_destroy(data[i]->iter);
        free(data[i]);
    }
    free(data); free(plp); free(ref); free(n_plp);
    return ret;
}
Esempio n. 9
0
int main_bedcov(int argc, char *argv[])
{
    gzFile fp;
    kstring_t str;
    kstream_t *ks;
    hts_idx_t **idx;
    aux_t **aux;
    int *n_plp, dret, i, n, c, min_mapQ = 0;
    int64_t *cnt;
    const bam_pileup1_t **plp;

    while ((c = getopt(argc, argv, "Q:")) >= 0) {
        switch (c) {
        case 'Q': min_mapQ = atoi(optarg); break;
        }
    }
    if (optind + 2 > argc) {
        fprintf(stderr, "Usage: samtools bedcov <in.bed> <in1.bam> [...]\n");
        return 1;
    }
    memset(&str, 0, sizeof(kstring_t));
    n = argc - optind - 1;
    aux = calloc(n, sizeof(aux_t*));
    idx = calloc(n, sizeof(hts_idx_t*));
    for (i = 0; i < n; ++i) {
        aux[i] = calloc(1, sizeof(aux_t));
        aux[i]->min_mapQ = min_mapQ;
        aux[i]->fp = sam_open(argv[i+optind+1], "r");
        idx[i] = sam_index_load(aux[i]->fp, argv[i+optind+1]);
        if (aux[i]->fp == 0 || idx[i] == 0) {
            fprintf(stderr, "ERROR: fail to open index BAM file '%s'\n", argv[i+optind+1]);
            return 2;
        }
        // TODO bgzf_set_cache_size(aux[i]->fp, 20);
        aux[i]->header = sam_hdr_read(aux[i]->fp);
    }
    cnt = calloc(n, 8);

    fp = gzopen(argv[optind], "rb");
    ks = ks_init(fp);
    n_plp = calloc(n, sizeof(int));
    plp = calloc(n, sizeof(bam_pileup1_t*));
    while (ks_getuntil(ks, KS_SEP_LINE, &str, &dret) >= 0) {
        char *p, *q;
        int tid, beg, end, pos;
        bam_mplp_t mplp;

        for (p = q = str.s; *p && *p != '\t'; ++p);
        if (*p != '\t') goto bed_error;
        *p = 0; tid = bam_name2id(aux[0]->header, q); *p = '\t';
        if (tid < 0) goto bed_error;
        for (q = p = p + 1; isdigit(*p); ++p);
        if (*p != '\t') goto bed_error;
        *p = 0; beg = atoi(q); *p = '\t';
        for (q = p = p + 1; isdigit(*p); ++p);
        if (*p == '\t' || *p == 0) {
            int c = *p;
            *p = 0; end = atoi(q); *p = c;
        } else goto bed_error;

        for (i = 0; i < n; ++i) {
            if (aux[i]->iter) hts_itr_destroy(aux[i]->iter);
            aux[i]->iter = sam_itr_queryi(idx[i], tid, beg, end);
        }
        mplp = bam_mplp_init(n, read_bam, (void**)aux);
        bam_mplp_set_maxcnt(mplp, 64000);
        memset(cnt, 0, 8 * n);
        while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0)
            if (pos >= beg && pos < end)
                for (i = 0; i < n; ++i) cnt[i] += n_plp[i];
        for (i = 0; i < n; ++i) {
            kputc('\t', &str);
            kputl(cnt[i], &str);
        }
        puts(str.s);
        bam_mplp_destroy(mplp);
        continue;

bed_error:
        fprintf(stderr, "Errors in BED line '%s'\n", str.s);
    }
    free(n_plp); free(plp);
    ks_destroy(ks);
    gzclose(fp);

    free(cnt);
    for (i = 0; i < n; ++i) {
        if (aux[i]->iter) hts_itr_destroy(aux[i]->iter);
        hts_idx_destroy(idx[i]);
        bam_hdr_destroy(aux[i]->header);
        sam_close(aux[i]->fp);
        free(aux[i]);
    }
    free(aux); free(idx);
    free(str.s);
    return 0;
}
Esempio n. 10
0
int main_depth(int argc, char *argv[])
{
    int i, n, tid, beg, end, pos, *n_plp, baseQ = 0, mapQ = 0, min_len = 0, status = EXIT_SUCCESS, nfiles;
    const bam_pileup1_t **plp;
    char *reg = 0; // specified region
    void *bed = 0; // BED data structure
    char *file_list = NULL, **fn = NULL;
    bam_hdr_t *h = NULL; // BAM header of the 1st input
    aux_t **data;
    bam_mplp_t mplp;

    // parse the command line
    while ((n = getopt(argc, argv, "r:b:q:Q:l:f:")) >= 0) {
        switch (n) {
            case 'l': min_len = atoi(optarg); break; // minimum query length
            case 'r': reg = strdup(optarg); break;   // parsing a region requires a BAM header
            case 'b':
                bed = bed_read(optarg); // BED or position list file can be parsed now
                if (!bed) { print_error_errno("Could not read file \"%s\"", optarg); return 1; }
                break;
            case 'q': baseQ = atoi(optarg); break;   // base quality threshold
            case 'Q': mapQ = atoi(optarg); break;    // mapping quality threshold
            case 'f': file_list = optarg; break;
        }
    }
    if (optind == argc && !file_list) {
        fprintf(pysamerr, "\n");
        fprintf(pysamerr, "Usage: samtools depth [options] in1.bam [in2.bam [...]]\n");
        fprintf(pysamerr, "Options:\n");
        fprintf(pysamerr, "   -b <bed>            list of positions or regions\n");
        fprintf(pysamerr, "   -f <list>           list of input BAM filenames, one per line [null]\n");
        fprintf(pysamerr, "   -l <int>            read length threshold (ignore reads shorter than <int>)\n");
        fprintf(pysamerr, "   -q <int>            base quality threshold\n");
        fprintf(pysamerr, "   -Q <int>            mapping quality threshold\n");
        fprintf(pysamerr, "   -r <chr:from-to>    region\n");
        fprintf(pysamerr, "\n");
        return 1;
    }

    // initialize the auxiliary data structures
    if (file_list)
    {
        if ( read_file_list(file_list,&nfiles,&fn) ) return 1;
        n = nfiles;
        argv = fn;
        optind = 0;
    }
    else
        n = argc - optind; // the number of BAMs on the command line
    data = calloc(n, sizeof(aux_t*)); // data[i] for the i-th input
    beg = 0; end = 1<<30;  // set the default region
    for (i = 0; i < n; ++i) {
        data[i] = calloc(1, sizeof(aux_t));
        data[i]->fp = sam_open(argv[optind+i], "r"); // open BAM
        if (data[i]->fp == NULL) {
            print_error_errno("Could not open \"%s\"", argv[optind+i]);
            status = EXIT_FAILURE;
            goto depth_end;
        }
        if (hts_set_opt(data[i]->fp, CRAM_OPT_REQUIRED_FIELDS,
                        SAM_FLAG | SAM_RNAME | SAM_POS | SAM_MAPQ | SAM_CIGAR |
                        SAM_SEQ)) {
            fprintf(pysamerr, "Failed to set CRAM_OPT_REQUIRED_FIELDS value\n");
            return 1;
        }
        if (hts_set_opt(data[i]->fp, CRAM_OPT_DECODE_MD, 0)) {
            fprintf(pysamerr, "Failed to set CRAM_OPT_DECODE_MD value\n");
            return 1;
        }
        data[i]->min_mapQ = mapQ;                    // set the mapQ filter
        data[i]->min_len  = min_len;                 // set the qlen filter
        data[i]->hdr = sam_hdr_read(data[i]->fp);    // read the BAM header
        if (reg) { // if a region is specified
            hts_idx_t *idx = sam_index_load(data[i]->fp, argv[optind+i]);  // load the index
            if (idx == NULL) {
                print_error("can't load index for \"%s\"", argv[optind+i]);
                status = EXIT_FAILURE;
                goto depth_end;
            }
            data[i]->iter = sam_itr_querys(idx, data[i]->hdr, reg); // set the iterator
            hts_idx_destroy(idx); // the index is not needed any more; free the memory
            if (data[i]->iter == NULL) {
                print_error("can't parse region \"%s\"", reg);
                status = EXIT_FAILURE;
                goto depth_end;
            }
        }
    }

    h = data[0]->hdr; // easy access to the header of the 1st BAM
    if (reg) {
        beg = data[0]->iter->beg; // and to the parsed region coordinates
        end = data[0]->iter->end;
    }

    // the core multi-pileup loop
    mplp = bam_mplp_init(n, read_bam, (void**)data); // initialization
    n_plp = calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM
    plp = calloc(n, sizeof(bam_pileup1_t*)); // plp[i] points to the array of covering reads (internal in mplp)
    while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0) { // come to the next covered position
        if (pos < beg || pos >= end) continue; // out of range; skip
        if (bed && bed_overlap(bed, h->target_name[tid], pos, pos + 1) == 0) continue; // not in BED; skip
        fputs(h->target_name[tid], stdout); printf("\t%d", pos+1); // a customized printf() would be faster
        for (i = 0; i < n; ++i) { // base level filters have to go here
            int j, m = 0;
            for (j = 0; j < n_plp[i]; ++j) {
                const bam_pileup1_t *p = plp[i] + j; // DON'T modfity plp[][] unless you really know
                if (p->is_del || p->is_refskip) ++m; // having dels or refskips at tid:pos
                else if (bam_get_qual(p->b)[p->qpos] < baseQ) ++m; // low base quality
            }
            printf("\t%d", n_plp[i] - m); // this the depth to output
        }
        putchar('\n');
    }
    free(n_plp); free(plp);
    bam_mplp_destroy(mplp);

depth_end:
    for (i = 0; i < n && data[i]; ++i) {
        bam_hdr_destroy(data[i]->hdr);
        if (data[i]->fp) sam_close(data[i]->fp);
        hts_itr_destroy(data[i]->iter);
        free(data[i]);
    }
    free(data); free(reg);
    if (bed) bed_destroy(bed);
    if ( file_list )
    {
        for (i=0; i<n; i++) free(fn[i]);
        free(fn);
    }
    return status;
}
Esempio n. 11
0
int bam2depth(const std::string& chromosomeName, const int startPos, const int endPos, const int minBaseQuality, const int minMappingQuality, const std::vector <std::string> & listOfFiles,
    std::vector< double > & averageCoveragePerBam
)
{
	int i, n, tid, beg, end, pos, *n_plp, baseQ = 0, mapQ = 0;
	const bam_pileup1_t **plp;
	char *reg = 0; // specified region
	//void *bed = 0; // BED data structure
	aux_t **data;
	bam_mplp_t mplp;

	// initialize the auxiliary data structures
	n = listOfFiles.size(); // the number of BAMs on the command line
	data = (aux_t **)calloc(n, sizeof(void*)); // data[i] for the i-th input
	beg = 0; end = 1<<30; tid = -1;  // set the default region
	beg = startPos;
	end = endPos;
	for (i = 0; i < n; ++i) {
		data[i] = (aux_t*)calloc(1, sizeof(aux_t));
		data[i]->fp = sam_open(listOfFiles[i].c_str(), "r"); // open BAM
		data[i]->min_mapQ = mapQ;                    // set the mapQ filter
		data[i]->hdr = sam_hdr_read(data[i]->fp);    // read the BAM header
		tid = bam_name2id(data[i]->hdr, chromosomeName.c_str());
		if (tid >= 0) { // if a region is specified and parsed successfully
			hts_idx_t *idx = sam_index_load(data[i]->fp, listOfFiles[i].c_str());  // load the index
			data[i]->iter = sam_itr_queryi(idx, tid, beg, end); // set the iterator
			hts_idx_destroy(idx); // the index is not needed any more; phase out of the memory
		}
	}

	// the core multi-pileup loop
	mplp = bam_mplp_init(n, read_bam, (void**)data); // initialization
	n_plp = (int*)calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM 
	plp = (const bam_pileup1_t **)calloc(n, sizeof(void*)); // plp[i] points to the array of covering reads (internal in mplp)
   std::vector<int> sumOfReadDepths( n, 0);
	while (bam_mplp_auto(mplp, &tid, &pos, n_plp, plp) > 0) { // come to the next covered position
		if (pos < beg || pos >= end) continue; // out of range; skip
		for (i = 0; i < n; ++i) { // base level filters have to go here
			int j, m = 0;
			for (j = 0; j < n_plp[i]; ++j) {
				const bam_pileup1_t *p = plp[i] + j; // DON'T modfity plp[][] unless you really know
				if (p->is_del || p->is_refskip) ++m; // having dels or refskips at tid:pos
				else if (bam_get_qual(p->b)[p->qpos] < baseQ) ++m; // low base quality
			}
			sumOfReadDepths[ i ] += n_plp[i] - m;
		}
	}
  	averageCoveragePerBam.resize( n );
    for (int fileIndex=0; fileIndex< n; fileIndex++ ) {
        averageCoveragePerBam[ fileIndex ] = (double)sumOfReadDepths[ fileIndex ] / (end - beg );  
    }
	free(n_plp); free(plp);
	bam_mplp_destroy(mplp);

	for (i = 0; i < n; ++i) {
		bam_hdr_destroy(data[i]->hdr);
		sam_close(data[i]->fp);
		if (data[i]->iter) hts_itr_destroy(data[i]->iter);
		free(data[i]);
	}
	free(data); free(reg);
	return 0;
}
Esempio n. 12
0
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, tid0 = -1, beg0 = 0, end0 = 1u<<29, ref_len, ref_tid = -1, max_depth, max_indel_depth;
	const bam_pileup1_t **plp;
	bam_mplp_t iter;
	bam_header_t *h = 0;
	char *ref;
	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 = 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], (conf->flag&MPLP_IGNORE_RG)? 0 : 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;
			idx = bam_index_load(fn[i]);
			if (idx == 0) {
				fprintf(stderr, "[%s] fail to load index for %d-th input.\n", __func__, i+1);
				exit(1);
			}
			if (bam_parse_region(h_tmp, conf->reg, &tid, &beg, &end) < 0) {
				fprintf(stderr, "[%s] malformatted region or wrong seqname for %d-th input.\n", __func__, i+1);
				exit(1);
			}
			if (i == 0) tid0 = tid, beg0 = beg, end0 = end;
			data[i]->iter = bam_iter_query(idx, tid, beg, end);
			bam_index_destroy(idx);
		}
		if (i == 0) h = h_tmp;
		else {
			// FIXME: to check consistency
			bam_header_destroy(h_tmp);
		}
	}
	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;
	}
	if (tid0 >= 0 && conf->fai) { // region is set
		ref = faidx_fetch_seq(conf->fai, h->target_name[tid0], 0, 0x7fffffff, &ref_len);
		ref_tid = tid0;
		for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid0;
	} else 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);


	int storeSize = 100;

	int delStore[2][100] = {{0},{0}};

	typedef char * mstring;

	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); ref = 0;
			if (conf->fai) ref = faidx_fetch_seq(conf->fai, h->target_name[tid], 0, 0x7fffffff, &ref_len);
			for (i = 0; i < n; ++i) data[i]->ref = ref, data[i]->ref_id = tid;
			ref_tid = tid;
		}
		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, conf->flag & MPLP_IGNORE_RG);
			_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*"); // FIXME: printf() is very slow...
					if (conf->flag & MPLP_PRINT_POS) printf("\t*");
				} else {
					//MDW start					
					//for each position in the pileup column
					int charLen = 16;
					int countChars[ charLen ][2];
					int countiChars[ charLen ][2];

					int countGap[2]={0,0};

					//double qvTotal=0;
					int numStruck=0;
					int numGood=0;
					int tti;
					int ttj;
					mstring insAllele[100];
					int insAlleleCnt[100];
					int sf=0;
					int flag=0;

					//typedef char * string;
					char insStr0[10000];
					int iCnt0=0;

					char insStr1[10000];
					int iCnt1=0;

					char delStr0[10000];
					int dCnt0=0;

					char delStr1[10000];
					int dCnt1=0;


					float qposP[10000];
					int qposCnt=0;



					//initialize with zeros
						for(tti=0;tti<charLen;tti++){
						  countChars[tti][0]=0;
						  countChars[tti][1]=0;
						}

					// define repeat length here; look back up to 10 prior positions
					// start one position away.
					int replC=0; //
					for(tti=1;tti<=15;tti++){
						// check for greater than zero
						if(toupper(ref[pos-1])==toupper(ref[pos-tti])){
							replC++;
						}else{ // breaks the chain at first non identical to current position not strict homopolymer
							break;
						}
					}					
					int reprC=0; // 
					for(tti=1;tti<=15;tti++){
						// check for greater than zero
						if(toupper(ref[pos+1])==toupper(ref[pos+tti])){
							reprC++;
						}else{ // breaks the chain at first non identical to current position not strict homopolymer
							break;
						}
					}		
					int repT = replC;
					if(replC < reprC){
						repT=reprC;
					}



					for (j = 0; j < n_plp[i]; ++j){
						const bam_pileup1_t *p = plp[i] + j;
									
						/*
						SAME LOGIC AS pileup_seq()
						*/

						if(p->is_refskip){ // never count intron gaps in numStruck
							continue;
						}

						if(p->is_del){ // skip deletion gap, after first position which is the first aligned char
							continue;
						}

						if( 	p->b->core.qual < conf->min_mqToCount  || // mapping quality
							conf->maxrepC < (repT) || // max homopolymer run, this will not 
							(!p->is_del && bam1_qual(p->b)[p->qpos] < conf->min_baseQ) || // base quality for matches
							p->alignedQPosBeg <= (conf->trimEnd ) || p->alignedQPosEnd <= (conf->trimEnd ) ||  // trimEnd is 1-based
							p->zf == 1 || // fusion tag
							p->ih > conf->maxIH  || // max hit index
							(p->nmd > conf->maxNM) || // max mismatch
							(conf->flagFilter == 1 && !(p->b->core.flag&BAM_FPROPER_PAIR)) || // optionally keep only proper pairs
							(conf->flagFilter == 2 && p->b->core.flag&BAM_FSECONDARY) || // optionally strike secondary
							(conf->flagFilter == 3 && p->b->core.flag&BAM_FDUP) || // optionally strike dup
							(conf->flagFilter == 4 && (p->b->core.flag&BAM_FDUP || p->b->core.flag&BAM_FSECONDARY))  || // optionally strike secondary or dup
							(conf->flagFilter == 5 && (p->b->core.flag&BAM_FDUP || p->b->core.flag&BAM_FSECONDARY || p->b->core.flag&BAM_FQCFAIL || !(p->b->core.flag&BAM_FPROPER_PAIR) ))   // optionally strike secondary, dup and QCfail


						){
							numStruck++;
							continue;
						}

						
						//printf("repT=%d: %d %c %c %c %c \n",repT,p->indel,ref[pos],ref[pos-1],ref[pos-2],ref[pos-3]);


						if(!p->is_del && p->indel==0){
  						  countChars[ bam1_seqi(bam1_seq(p->b), p->qpos) ][ bam1_strand(p->b) ] ++;
						  numGood++;			

						}else if(p->is_refskip){
						  countGap[ bam1_strand(p->b) ]++;
						}
						
						if(p->indel<0){
    						  numGood++;			
						  if(bam1_strand(p->b) ==0){
							  for(tti=1;tti<= -p->indel; tti++) {
							    // current spot, starting at 0 in store, because indel<0 refers to next position
							   delStr0[dCnt0] =  ref[pos+tti];
							   dCnt0++;
							  }	
							  delStr0[dCnt0] = ',';
							  dCnt0++;
						  }else{
							  for(tti=1;tti<= -p->indel; tti++) {
							    // current spot, starting at 0 in store, because indel<0 refers to next position
							   delStr1[dCnt1] = ref[pos+tti];
							   dCnt1++;
							  }	
							  delStr1[dCnt1] = ',';
							  dCnt1++;
						  }



						}else if(p->indel>0){
						  numGood++;			

						  if(bam1_strand(p->b) ==0){
							  for(tti=1;tti<= p->indel; tti++) {
							    // current spot, starting at 0 in store, because indel<0 refers to next position
							   insStr0[iCnt0] = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos + tti)];
							   iCnt0++;
							  }	
							  insStr0[iCnt0] = ',';
							  iCnt0++;
						  }else{
							  for(tti=1;tti<= p->indel; tti++) {
							    // current spot, starting at 0 in store, because indel<0 refers to next position
							   insStr1[iCnt1] = bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos + tti)];
							   iCnt1++;
							  }	
							  insStr1[iCnt1] = ',';
							  iCnt1++;
						  }


						}
						//calculate position of variant within aligned read - no soft clips
						if( toupper(ref[pos]) != toupper(bam_nt16_rev_table[bam1_seqi(bam1_seq(p->b), p->qpos)]) || p->indel>0 || p->indel<0  ){

						//distance to end; calculate distance to end of aligned read.  removes soft clips.
						int distToEnd = (p->alignedQPosBeg < p->alignedQPosEnd) ? p->alignedQPosBeg : p->alignedQPosEnd;
						qposP[qposCnt] = distToEnd;						  
						qposCnt++;	
						// printf("id=%s, pos=%d",bam1_qname(p->b),distToEnd);
						}	
					}

					//

					//print A,C,G,T, by +/-
				        printf("\t%d\t%d\t%d\t%d\t%d\t%d\t%d\t%d\t%d\t%d", 	countChars[1][0],countChars[1][1],
											countChars[2][0],countChars[2][1],
											countChars[4][0],countChars[4][1],
											countChars[8][0],countChars[8][1],
											countChars[7][0],countChars[7][1]);
					
					putchar('\t');
					for(tti=0;tti<dCnt0;tti++){
					  putchar(delStr0[tti]);
					}

					putchar('\t');
					for(tti=0;tti<dCnt1;tti++){
					  putchar(delStr1[tti]);
					}

					putchar('\t');
					for(tti=0;tti<iCnt0;tti++){
					  putchar(insStr0[tti]);
					}

					putchar('\t');
					for(tti=0;tti<iCnt1;tti++){
					  putchar(insStr1[tti]);
					}

					printf("\t%d\t%d",numGood,numStruck);					

					// get non-ref qpos variation

					float medqpos = -1;
					float medAbsDev = -1;
					if(qposCnt>0){
					  medqpos = median(qposCnt,qposP);
					  float absDev[qposCnt];
					  for(tti=0;tti<qposCnt;tti++){
						absDev[tti] = abs(medqpos - qposP[tti]);
					  }
					  medAbsDev = median(qposCnt-1,absDev);
					}
					printf("\t%f",medAbsDev);

					///END MDW
				}



			}
			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) {
		bam_close(data[i]->fp);
		if (data[i]->iter) bam_iter_destroy(data[i]->iter);
		free(data[i]);
	}
	free(data); free(plp); free(ref); free(n_plp);
	return 0;
}
Esempio n. 13
0
void extractCalls(Config *config) {
    bam_hdr_t *hdr = sam_hdr_read(config->fp);
    bam_mplp_t iter;
    int ret, tid, pos, i, seqlen, type, rv, o = 0;
    int beg0 = 0, end0 = 1u<<29;
    int n_plp; //This will need to be modified for multiple input files
    int ctid = -1; //The tid of the contig whose sequence is stored in "seq"
    int idxBED = 0, strand;
    uint32_t nmethyl = 0, nunmethyl = 0;
    const bam_pileup1_t **plp = NULL;
    char *seq = NULL, base;
    mplp_data *data = NULL;
    struct lastCall *lastCpG = NULL;
    struct lastCall *lastCHG = NULL;

    if(config->merge) {
        if(config->keepCpG) {
            lastCpG = calloc(1, sizeof(struct lastCall));
            assert(lastCpG);
            lastCpG->tid = -1;
        }
        if(config->keepCHG) {
            lastCHG = calloc(1, sizeof(struct lastCall));
            assert(lastCHG);
            lastCHG->tid = -1;
        }
    }

    data = calloc(1,sizeof(mplp_data));
    if(data == NULL) {
        fprintf(stderr, "Couldn't allocate space for the data structure in extractCalls()!\n");
        return;
    }
    data->config = config;
    data->hdr = hdr;
    if (config->reg) {
        if((data->iter = sam_itr_querys(config->bai, hdr, config->reg)) == 0) {
            fprintf(stderr, "failed to parse regions %s", config->reg);
            return;
        }
    }
    if(config->bedName) {
        config->bed = parseBED(config->bedName, hdr);
        if(config->bed == NULL) return;
    }

    plp = calloc(1, sizeof(bam_pileup1_t *)); //This will have to be modified for multiple input files
    if(plp == NULL) {
        fprintf(stderr, "Couldn't allocate space for the plp structure in extractCalls()!\n");
        return;
    }

    //Start the pileup
    iter = bam_mplp_init(1, filter_func, (void **) &data);
    bam_mplp_init_overlaps(iter);
    bam_mplp_set_maxcnt(iter, config->maxDepth);
    while((ret = cust_mplp_auto(iter, &tid, &pos, &n_plp, plp)) > 0) {
        //Do we need to process this position?
        if (config->reg) {
            beg0 = data->iter->beg, end0 = data->iter->end;
            if ((pos < beg0 || pos >= end0)) continue; // out of the region requested
        }
        if(tid != ctid) {
            if(seq != NULL) free(seq);
            seq = faidx_fetch_seq(config->fai, hdr->target_name[tid], 0, faidx_seq_len(config->fai, hdr->target_name[tid]), &seqlen);
            if(seqlen < 0) {
                fprintf(stderr, "faidx_fetch_seq returned %i while trying to fetch the sequence for tid %i (%s)!\n",\
                        seqlen, tid, hdr->target_name[tid]);
                fprintf(stderr, "Note that the output will be truncated!\n");
                return;
            }
            ctid = tid;
        }

        if(config->bed) { //Handle -l
            while((o = posOverlapsBED(tid, pos, config->bed, idxBED)) == -1) idxBED++;
            if(o == 0) continue; //Wrong strand
        }

        if(isCpG(seq, pos, seqlen)) {
            if(!config->keepCpG) continue;
            type = 0;
        } else if(isCHG(seq, pos, seqlen)) {
            if(!config->keepCHG) continue;
            type = 1;
        } else if(isCHH(seq, pos, seqlen)) {
            if(!config->keepCHH) continue;
            type = 2;
        } else {
            continue;
        }

        nmethyl = nunmethyl = 0;
        base = *(seq+pos);
        for(i=0; i<n_plp; i++) {
            if(plp[0][i].is_del) continue;
            if(plp[0][i].is_refskip) continue;
            if(config->bed) if(!readStrandOverlapsBED(plp[0][i].b, config->bed->region[idxBED])) continue;
            strand = getStrand((plp[0]+i)->b);
            if(strand & 1) {
                if(base != 'C' && base != 'c') continue;
            } else {
                if(base != 'G' && base != 'g') continue;
            }
            rv = updateMetrics(config, plp[0]+i);
            if(rv > 0) nmethyl++;
            else if(rv<0) nunmethyl++;
        }

        if(nmethyl+nunmethyl==0) continue;
        if(!config->merge || type==2) {
            writeCall(config->output_fp[type], config, hdr->target_name[tid], pos, 1, nmethyl, nunmethyl);
        } else {
            //Merge into per-CpG/CHG metrics
            if(type==0) {
                if(base=='G' || base=='g') pos--;
                processLast(config->output_fp[0], config, lastCpG, hdr, tid, pos, 2, nmethyl, nunmethyl);
            } else {
                if(base=='G' || base=='g') pos-=2;
                processLast(config->output_fp[1], config, lastCHG, hdr, tid, pos, 3, nmethyl, nunmethyl);
            }
        }
    }

    //Don't forget the last CpG/CHG
    if(config->merge) {
        if(config->keepCpG && lastCpG->tid != -1) {
            processLast(config->output_fp[0], config, lastCpG, hdr, tid, pos, 2, nmethyl, nunmethyl);
        }
        if(config->keepCHG && lastCHG->tid != -1) {
            processLast(config->output_fp[1], config, lastCHG, hdr, tid, pos, 3, nmethyl, nunmethyl);
        }
    }

    bam_hdr_destroy(hdr);
    if(data->iter) hts_itr_destroy(data->iter);
    bam_mplp_destroy(iter);
    free(data);
    free(plp);
    if(seq != NULL) free(seq);
}
Esempio n. 14
0
int main(int argc, char *argv[])
{
    int c, i, n, ret, res;
    int tid, pos, *n_plp;
    cmdopt_t o;
    bam_mplp_t mplp;
    const bam_pileup1_t **plp;
    aux_t **data;
    bam_hdr_t *h = 0;
    sv_t sv1;
    qual_sum_t qual2;
    khiter_t k_iter;
    khash_t(sv_hash) *sv_h = kh_init(sv_hash);
    khash_t(sv_geno) *geno_h = kh_init(sv_geno);
    khash_t(colmap) *smp_cols;
    khash_t(ped) *ped_h = 0;
    mempool_t *mp;
    char **samples;
    
    o.min_q = 40; o.min_s = 80; o.min_len = 150; o.min_dp = 10; o.bed = 0, o.fnped = 0, o.mi_prob=0.005;
    while ((c = getopt(argc, argv, "hq:s:l:d:b:p:m:")) >= 0) {
        if (c == 'h') { usage(stderr, &o); return 0; }
        else if (c == 'q') o.min_q = atoi(optarg);
        else if (c == 's') o.min_s = atoi(optarg);
        else if (c == 'l') o.min_len = atoi(optarg);
        else if (c == 'd') o.min_dp = atoi(optarg);
        else if (c == 'p') o.fnped = optarg;
        else if (c == 'm') o.mi_prob = atof(optarg);
        else if (c == 'b') { 
            if ((o.bed = bed_read(optarg)) == NULL) {
                return -1;
            }
        }
    }
    if (o.mi_prob < 0.0000000000001 || o.mi_prob > 0.1) {
        fprintf(stderr, "Error. Probability of a mendelian inconsistency must be between 0.1 and 0.0000000000001.\n");
    }
    
    if (argc - optind < 1) {
        usage(stderr, &o);
        return 1;
    }

    // Open files and initalize aux data //
    n = argc - optind;
    data = calloc(n, sizeof(aux_t*));
    samples = (char**)malloc(n * sizeof(char*));
    for (i = 0; i < n; ++i) {
        data[i] = calloc(1, sizeof (aux_t));
        data[i]->fp = sam_open(argv[optind + i], "r");
        if (!data[i]->fp) {
            fprintf(stderr, "Input file \"%s\" could not be opened.\n", argv[optind + 1]);
            return 1;
        }
        data[i]->min_mapq = o.min_q;
        data[i]->min_as = o.min_s;
        data[i]->min_len = o.min_len;
        data[i]->hdr = sam_hdr_read(data[i]->fp);
        if (!data[i]->hdr) {
            fprintf(stderr, "Could not read the header for input file \"%s\".\n", argv[optind + 1]);
            return 1;
        }
        samples[i] = find_sample(data[i]->hdr, &res);
        if (!samples[i]) {
            fprintf(stderr, "Warning. No sample name detected for bam %s. Using filename\n", argv[optind + i]);
            samples[i] = argv[optind + i];
        }
    }
    h = data[0]->hdr;
    smp_cols = map_samples(samples, n);
    if (o.fnped) {
        if ((ped_h = read_ped(o.fnped, smp_cols)) == 0) { return -1; }
    }

    // The core data processing loop //
    mplp = bam_mplp_init(n, read_bam, (void**)data);
    n_plp = calloc(n, sizeof(int)); // n_plp[i] is the number of covering reads from the i-th BAM
    plp = calloc(n, sizeof(bam_pileup1_t*)); // plp[i] points to the array of covering reads in mplp
    //quals = (qual_vec_t*)calloc(n, sizeof(qual_vec_t));
    mp = mp_init();
    while ((ret = bam_mplp_auto(mplp, &tid, &pos, n_plp, plp)) > 0) { // iterate of positions with coverage
        int n_sv;
        if (o.bed && tid >= 0 && !bed_overlap(o.bed, h->target_name[tid], pos, pos+1)) continue;
        n_sv = plp2sv(h, tid, pos, n, n_plp, plp, sv_h);
        if (n_sv > 1) { fprintf(stderr, "Warning: more than two alleles detected at %s:%d\n", h->target_name[tid], pos); }
        if (n_sv) {
            fprintf(stderr, "SV detected at %d:%d\n", tid, pos);
            for (k_iter = kh_begin(sv_h); k_iter != kh_end(sv_h); ++k_iter) {
                if (kh_exist(sv_h, k_iter)) {
                    sv1 = kh_value(sv_h, k_iter);
                    fprintf(stderr, "SV tid1=%d, tid2=%d, pos1=%d, pos2=%d, ori1=%d, ori2=%d, allele=%d\n", sv1.tid1, sv1.tid2, sv1.pos1, sv1.pos2, sv1.ori1, sv1.ori2, sv1.allele);
                }
            }
            res = get_qual_data(h, tid, pos, n, n_plp, plp, n_sv + 1, sv_h, geno_h, mp);
            if (res < 0) {
                fprintf(stderr, "Error collecting quality data from reads\n");
                return -1;
            }
            kh_clear(sv_hash, sv_h);
        }
    }

    print_header(h, optind, n, argv);
    genotype_sv(h, n, geno_h, o.min_dp, ped_h, o.mi_prob);

    free(n_plp);
    free(plp);
    bam_mplp_destroy(mplp);
    mp_destroy(mp);
    if (o.bed) bed_destroy(o.bed);
    for (i = 0; i < n; ++i) { 
        bam_hdr_destroy(data[i]->hdr);
        sam_close(data[i]->fp);
        free(data[i]);
        free(samples[i]);
    }
    free(data);
    free(samples);
    kh_destroy(sv_hash, sv_h);
    kh_destroy(sv_geno, geno_h);
    kh_destroy(colmap, smp_cols);
    kh_destroy(ped, ped_h);
    return 0;
}