/* bam_md.c -- calmd subcommand. Copyright (C) 2009-2011, 2014-2015 Genome Research Ltd. Portions copyright (C) 2009-2011 Broad Institute. Author: Heng Li Permission is hereby granted, free of charge, to any person obtaining a copy of this software and associated documentation files (the "Software"), to deal in the Software without restriction, including without limitation the rights to use, copy, modify, merge, publish, distribute, sublicense, and/or sell copies of the Software, and to permit persons to whom the Software is furnished to do so, subject to the following conditions: The above copyright notice and this permission notice shall be included in all copies or substantial portions of the Software. THE SOFTWARE IS PROVIDED "AS IS", WITHOUT WARRANTY OF ANY KIND, EXPRESS OR IMPLIED, INCLUDING BUT NOT LIMITED TO THE WARRANTIES OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE AND NONINFRINGEMENT. IN NO EVENT SHALL THE AUTHORS OR COPYRIGHT HOLDERS BE LIABLE FOR ANY CLAIM, DAMAGES OR OTHER LIABILITY, WHETHER IN AN ACTION OF CONTRACT, TORT OR OTHERWISE, ARISING FROM, OUT OF OR IN CONNECTION WITH THE SOFTWARE OR THE USE OR OTHER DEALINGS IN THE SOFTWARE. */ #include #include #include #include #include #include #include "htslib/faidx.h" #include "htslib/sam.h" #include "htslib/kstring.h" #include "htslib/thread_pool.h" #include "sam_opts.h" #include "samtools.h" #define USE_EQUAL 1 #define DROP_TAG 2 #define BIN_QUAL 4 #define UPDATE_NM 8 #define UPDATE_MD 16 #define HASH_QNM 32 int bam_aux_drop_other(bam1_t *b, uint8_t *s); void bam_fillmd1_core(bam1_t *b, char *ref, int ref_len, int flag, int max_nm) { uint8_t *seq = bam_get_seq(b); uint32_t *cigar = bam_get_cigar(b); bam1_core_t *c = &b->core; int i, x, y, u = 0; kstring_t *str; int32_t old_nm_i = -1, nm = 0; str = (kstring_t*)calloc(1, sizeof(kstring_t)); for (i = y = 0, x = c->pos; i < c->n_cigar; ++i) { int j, l = cigar[i]>>4, op = cigar[i]&0xf; if (op == BAM_CMATCH || op == BAM_CEQUAL || op == BAM_CDIFF) { for (j = 0; j < l; ++j) { int c1, c2, z = y + j; if (x+j >= ref_len || ref[x+j] == '\0') break; // out of bounds c1 = bam_seqi(seq, z), c2 = seq_nt16_table[(int)ref[x+j]]; if ((c1 == c2 && c1 != 15 && c2 != 15) || c1 == 0) { // a match if (flag&USE_EQUAL) seq[z/2] &= (z&1)? 0xf0 : 0x0f; ++u; } else { kputw(u, str); kputc(ref[x+j], str); u = 0; ++nm; } } if (j < l) break; x += l; y += l; } else if (op == BAM_CDEL) { kputw(u, str); kputc('^', str); for (j = 0; j < l; ++j) { if (x+j >= ref_len || ref[x+j] == '\0') break; kputc(ref[x+j], str); } u = 0; x += j; nm += j; if (j < l) break; } else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) { y += l; if (op == BAM_CINS) nm += l; } else if (op == BAM_CREF_SKIP) { x += l; } } kputw(u, str); // apply max_nm if (max_nm > 0 && nm >= max_nm) { for (i = y = 0, x = c->pos; i < c->n_cigar; ++i) { int j, l = cigar[i]>>4, op = cigar[i]&0xf; if (op == BAM_CMATCH || op == BAM_CEQUAL || op == BAM_CDIFF) { for (j = 0; j < l; ++j) { int c1, c2, z = y + j; if (x+j >= ref_len || ref[x+j] == '\0') break; // out of bounds c1 = bam_seqi(seq, z), c2 = seq_nt16_table[(int)ref[x+j]]; if ((c1 == c2 && c1 != 15 && c2 != 15) || c1 == 0) { // a match seq[z/2] |= (z&1)? 0x0f : 0xf0; bam_get_qual(b)[z] = 0; } } if (j < l) break; x += l; y += l; } else if (op == BAM_CDEL || op == BAM_CREF_SKIP) x += l; else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) y += l; } } // update NM if ((flag & UPDATE_NM) && !(c->flag & BAM_FUNMAP)) { uint8_t *old_nm = bam_aux_get(b, "NM"); if (old_nm) old_nm_i = bam_aux2i(old_nm); if (!old_nm) bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm); else if (nm != old_nm_i) { fprintf(stderr, "[bam_fillmd1] different NM for read '%s': %d -> %d\n", bam_get_qname(b), old_nm_i, nm); bam_aux_del(b, old_nm); bam_aux_append(b, "NM", 'i', 4, (uint8_t*)&nm); } } // update MD if ((flag & UPDATE_MD) && !(c->flag & BAM_FUNMAP)) { uint8_t *old_md = bam_aux_get(b, "MD"); if (!old_md) bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s); else { int is_diff = 0; if (strlen((char*)old_md+1) == str->l) { for (i = 0; i < str->l; ++i) if (toupper(old_md[i+1]) != toupper(str->s[i])) break; if (i < str->l) is_diff = 1; } else is_diff = 1; if (is_diff) { fprintf(stderr, "[bam_fillmd1] different MD for read '%s': '%s' -> '%s'\n", bam_get_qname(b), old_md+1, str->s); bam_aux_del(b, old_md); bam_aux_append(b, "MD", 'Z', str->l + 1, (uint8_t*)str->s); } } } // drop all tags but RG if (flag&DROP_TAG) { uint8_t *q = bam_aux_get(b, "RG"); bam_aux_drop_other(b, q); } // reduce the resolution of base quality if (flag&BIN_QUAL) { uint8_t *qual = bam_get_qual(b); for (i = 0; i < b->core.l_qseq; ++i) if (qual[i] >= 3) qual[i] = qual[i]/10*10 + 7; } free(str->s); free(str); } void bam_fillmd1(bam1_t *b, char *ref, int flag) { bam_fillmd1_core(b, ref, INT_MAX, flag, 0); } int calmd_usage() { fprintf(stderr, "Usage: samtools calmd [-eubrAES] \n" "Options:\n" " -e change identical bases to '='\n" " -u uncompressed BAM output (for piping)\n" " -b compressed BAM output\n" " -S ignored (input format is auto-detected)\n" " -A modify the quality string\n" " -r compute the BQ tag (without -A) or cap baseQ by BAQ (with -A)\n" " -E extended BAQ for better sensitivity but lower specificity\n"); sam_global_opt_help(stderr, "-....@"); return 1; } int bam_fillmd(int argc, char *argv[]) { int c, flt_flag, tid = -2, ret, len, is_bam_out, is_uncompressed, max_nm, is_realn, capQ, baq_flag; htsThreadPool p = {NULL, 0}; samFile *fp = NULL, *fpout = NULL; bam_hdr_t *header = NULL; faidx_t *fai = NULL; char *ref = NULL, mode_w[8], *ref_file; bam1_t *b = NULL; sam_global_args ga = SAM_GLOBAL_ARGS_INIT; static const struct option lopts[] = { SAM_OPT_GLOBAL_OPTIONS('-', 0, 0, 0, 0,'@'), { NULL, 0, NULL, 0 } }; flt_flag = UPDATE_NM | UPDATE_MD; is_bam_out = is_uncompressed = is_realn = max_nm = capQ = baq_flag = 0; strcpy(mode_w, "w"); while ((c = getopt_long(argc, argv, "EqreuNhbSC:n:Ad@:", lopts, NULL)) >= 0) { switch (c) { case 'r': is_realn = 1; break; case 'e': flt_flag |= USE_EQUAL; break; case 'd': flt_flag |= DROP_TAG; break; case 'q': flt_flag |= BIN_QUAL; break; case 'h': flt_flag |= HASH_QNM; break; case 'N': flt_flag &= ~(UPDATE_MD|UPDATE_NM); break; case 'b': is_bam_out = 1; break; case 'u': is_uncompressed = is_bam_out = 1; break; case 'S': break; case 'n': max_nm = atoi(optarg); break; case 'C': capQ = atoi(optarg); break; case 'A': baq_flag |= 1; break; case 'E': baq_flag |= 2; break; default: if (parse_sam_global_opt(c, optarg, lopts, &ga) == 0) break; fprintf(stderr, "[bam_fillmd] unrecognized option '-%c'\n\n", c); /* else fall-through */ case '?': return calmd_usage(); } } if (is_bam_out) strcat(mode_w, "b"); else strcat(mode_w, "h"); if (is_uncompressed) strcat(mode_w, "0"); if (optind + (ga.reference == NULL) >= argc) return calmd_usage(); fp = sam_open_format(argv[optind], "r", &ga.in); if (fp == NULL) { print_error_errno("calmd", "Failed to open input file '%s'", argv[optind]); return 1; } header = sam_hdr_read(fp); if (header == NULL || header->n_targets == 0) { fprintf(stderr, "[bam_fillmd] input SAM does not have header. Abort!\n"); goto fail; } fpout = sam_open_format("-", mode_w, &ga.out); if (fpout == NULL) { print_error_errno("calmd", "Failed to open output"); goto fail; } if (sam_hdr_write(fpout, header) < 0) { print_error_errno("calmd", "Failed to write sam header"); goto fail; } if (ga.nthreads > 0) { if (!(p.pool = hts_tpool_init(ga.nthreads))) { fprintf(stderr, "Error creating thread pool\n"); goto fail; } hts_set_opt(fp, HTS_OPT_THREAD_POOL, &p); hts_set_opt(fpout, HTS_OPT_THREAD_POOL, &p); } ref_file = argc > optind + 1 ? argv[optind+1] : ga.reference; fai = fai_load(ref_file); if (!fai) { print_error_errno("calmd", "Failed to open reference file '%s'", ref_file); goto fail; } b = bam_init1(); if (!b) { fprintf(stderr, "[bam_fillmd] Failed to allocate bam struct\n"); goto fail; } while ((ret = sam_read1(fp, header, b)) >= 0) { if (b->core.tid >= 0) { if (tid != b->core.tid) { free(ref); ref = fai_fetch(fai, header->target_name[b->core.tid], &len); tid = b->core.tid; if (ref == 0) { // FIXME: Should this always be fatal? fprintf(stderr, "[bam_fillmd] fail to find sequence '%s' in the reference.\n", header->target_name[tid]); if (is_realn || capQ > 10) goto fail; // Would otherwise crash } } if (is_realn) sam_prob_realn(b, ref, len, baq_flag); if (capQ > 10) { int q = sam_cap_mapq(b, ref, len, capQ); if (b->core.qual > q) b->core.qual = q; } if (ref) bam_fillmd1_core(b, ref, len, flt_flag, max_nm); } if (sam_write1(fpout, header, b) < 0) { print_error_errno("calmd", "failed to write to output file"); goto fail; } } if (ret < -1) { fprintf(stderr, "[bam_fillmd] Error reading input.\n"); goto fail; } bam_destroy1(b); bam_hdr_destroy(header); free(ref); fai_destroy(fai); sam_close(fp); if (sam_close(fpout) < 0) { fprintf(stderr, "[bam_fillmd] error when closing output file\n"); return 1; } if (p.pool) hts_tpool_destroy(p.pool); return 0; fail: free(ref); if (b) bam_destroy1(b); if (header) bam_hdr_destroy(header); if (fai) fai_destroy(fai); if (fp) sam_close(fp); if (fpout) sam_close(fpout); if (p.pool) hts_tpool_destroy(p.pool); return 1; }