/* bam_sort.c -- sorting and merging. Copyright (C) 2008-2015 Genome Research Ltd. Portions copyright (C) 2009-2012 Broad Institute. Author: Heng Li Author: Martin Pollard 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 #include #include #include #include "htslib/ksort.h" #include "htslib/khash.h" #include "htslib/klist.h" #include "htslib/kstring.h" #include "htslib/sam.h" #include "sam_opts.h" #if !defined(__DARWIN_C_LEVEL) || __DARWIN_C_LEVEL < 900000L #define NEED_MEMSET_PATTERN4 #endif #ifdef NEED_MEMSET_PATTERN4 void memset_pattern4(void *target, const void *pattern, size_t size) { uint32_t* target_iter = target; size_t loops = size/4; size_t i; for (i = 0; i < loops; ++i) { memcpy(target_iter, pattern, 4); ++target_iter; } if (size%4 != 0) memcpy(target_iter, pattern, size%4); } #endif KHASH_INIT(c2c, char*, char*, 1, kh_str_hash_func, kh_str_hash_equal) KHASH_INIT(cset, char*, char, 0, kh_str_hash_func, kh_str_hash_equal) KHASH_MAP_INIT_STR(c2i, int) #define hdrln_free_char(p) KLIST_INIT(hdrln, char*, hdrln_free_char) static int g_is_by_qname = 0; // Added for RSEM by Bo Li: Test if the query name should stop static bool ok(char c) { return c > 0 && !isspace(c); } // Modified for RSEM by Bo Li: Let the query name stops whenever encounters a space static int strnum_cmp(const char *_a, const char *_b) { const unsigned char *a = (const unsigned char*)_a, *b = (const unsigned char*)_b; const unsigned char *pa = a, *pb = b; //while (*pa && *pb) { // Modified for RSEM by Bo Li while (ok(*pa) && ok(*pb)) { if (isdigit(*pa) && isdigit(*pb)) { while (*pa == '0') ++pa; while (*pb == '0') ++pb; while (isdigit(*pa) && isdigit(*pb) && *pa == *pb) ++pa, ++pb; if (isdigit(*pa) && isdigit(*pb)) { int i = 0; while (isdigit(pa[i]) && isdigit(pb[i])) ++i; return isdigit(pa[i])? 1 : isdigit(pb[i])? -1 : (int)*pa - (int)*pb; } else if (isdigit(*pa)) return 1; else if (isdigit(*pb)) return -1; else if (pa - a != pb - b) return pa - a < pb - b? 1 : -1; } else { if (*pa != *pb) return (int)*pa - (int)*pb; ++pa; ++pb; } } //return *pa ? 1 : *pb ? -1 : 0; // Modified for RSEM by Bo Li return ok(*pa)? 1 : ok(*pb)? -1 : 0; } #define HEAP_EMPTY UINT64_MAX typedef struct { int i; uint64_t pos, idx; bam1_t *b; } heap1_t; #define __pos_cmp(a, b) ((a).pos > (b).pos || ((a).pos == (b).pos && ((a).i > (b).i || ((a).i == (b).i && (a).idx > (b).idx)))) // Function to compare reads in the heap and determine which one is < the other // Note by Bo Li: It actually returns which one is > static inline int heap_lt(const heap1_t a, const heap1_t b) { if (g_is_by_qname) { int t; if (a.b == NULL || b.b == NULL) return a.b == NULL? 1 : 0; t = strnum_cmp(bam_get_qname(a.b), bam_get_qname(b.b)); //return (t > 0 || (t == 0 && (a.b->core.flag&0xc0) > (b.b->core.flag&0xc0))); // Modified for RSEM by Bo Li return t > 0; // In order to make sure the two mates of an alignment are adjacent to each other } else return __pos_cmp(a, b); } KSORT_INIT(heap, heap1_t, heap_lt) typedef struct merged_header { kstring_t out_hd; kstring_t out_sq; kstring_t out_rg; kstring_t out_pg; kstring_t out_co; char **target_name; uint32_t *target_len; size_t n_targets; size_t targets_sz; khash_t(c2i) *sq_tids; khash_t(cset) *rg_ids; khash_t(cset) *pg_ids; bool have_hd; } merged_header_t; typedef struct trans_tbl { int32_t n_targets; int* tid_trans; kh_c2c_t* rg_trans; kh_c2c_t* pg_trans; bool lost_coord_sort; } trans_tbl_t; /* Something to look like a regmatch_t */ typedef struct hdr_match { ptrdiff_t rm_so; ptrdiff_t rm_eo; } hdr_match_t; /* * Search for header lines of a particular record type. * * This replaces a regex search for something like /^@SQ.*\tSN:([^\t]+).*$/ * but is much quicker. The locations found are returned in *matches, * which has a signature the same as that of a regmatch_t. * * rec is the record type to match (i.e. @HD, @SQ, @PG or @RG) * tag is a tag type in the record to match (SN for @SQ, ID for @PG or @RG) * * The location of the record (if found) is returned in matches[0] * If tag is not NULL, the record is searched for the presence of the * given tag. If found, the location of the value is returned in matches[1]. * If the tag isn't found then the record is ignored and the search resumes * on the next header line. * * For simplicity, some assumptions are made about rec and tag: * rec should include the leading '@' sign and be three characters long. * tag should be exactly two characters long. * These are always string constants when this is called below, so we don't * bother to check here. * * Returns 0 if a match was found, -1 if not. */ static int hdr_line_match(const char *text, const char *rec, const char *tag, hdr_match_t *matches) { const char *line_start, *line_end = text; const char *tag_start, *tag_end; for (;;) { // Find record, ensure either at start of text or follows '\n' line_start = strstr(line_end, rec); while (line_start && line_start > text && *(line_start - 1) != '\n') { line_start = strstr(line_start + 3, rec); } if (!line_start) return -1; // Find end of header line line_end = strchr(line_start, '\n'); if (!line_end) line_end = line_start + strlen(line_start); matches[0].rm_so = line_start - text; matches[0].rm_eo = line_end - text; if (!tag) return 0; // Match found if not looking for tag. for (tag_start = line_start + 3; tag_start < line_end; tag_start++) { // Find possible tag start. Hacky but quick. while (*tag_start > '\n') tag_start++; // Check it if (tag_start[0] == '\t' && strncmp(tag_start + 1, tag, 2) == 0 && tag_start[3] == ':') { // Found tag, record location and return. tag_end = tag_start + 4; while (*tag_end && *tag_end != '\t' && *tag_end != '\n') ++tag_end; matches[1].rm_so = tag_start - text + 4; matches[1].rm_eo = tag_end - text; return 0; } } // Couldn't find tag, try again from end of current record. } } static void trans_tbl_destroy(trans_tbl_t *tbl) { khiter_t iter; free(tbl->tid_trans); /* * The values for the tbl->rg_trans and tbl->pg_trans hashes are pointers * to keys in the rg_ids and pg_ids sets of the merged_header_t, so * they should not be freed here. * * The keys are unique to each hash entry, so they do have to go. */ for (iter = kh_begin(tbl->rg_trans); iter != kh_end(tbl->rg_trans); ++iter) { if (kh_exist(tbl->rg_trans, iter)) { free(kh_key(tbl->rg_trans, iter)); } } for (iter = kh_begin(tbl->pg_trans); iter != kh_end(tbl->pg_trans); ++iter) { if (kh_exist(tbl->pg_trans, iter)) { free(kh_key(tbl->pg_trans, iter)); } } kh_destroy(c2c,tbl->rg_trans); kh_destroy(c2c,tbl->pg_trans); } /* * Create a merged_header_t struct. */ static merged_header_t * init_merged_header() { merged_header_t *merged_hdr; merged_hdr = calloc(1, sizeof(*merged_hdr)); if (merged_hdr == NULL) return NULL; merged_hdr->targets_sz = 16; merged_hdr->target_name = malloc(merged_hdr->targets_sz * sizeof(*merged_hdr->target_name)); if (NULL == merged_hdr->target_name) goto fail; merged_hdr->target_len = malloc(merged_hdr->targets_sz * sizeof(*merged_hdr->target_len)); if (NULL == merged_hdr->target_len) goto fail; merged_hdr->sq_tids = kh_init(c2i); if (merged_hdr->sq_tids == NULL) goto fail; merged_hdr->rg_ids = kh_init(cset); if (merged_hdr->rg_ids == NULL) goto fail; merged_hdr->pg_ids = kh_init(cset); if (merged_hdr->pg_ids == NULL) goto fail; return merged_hdr; fail: perror("[init_merged_header]"); kh_destroy(cset, merged_hdr->pg_ids); kh_destroy(cset, merged_hdr->rg_ids); kh_destroy(c2i, merged_hdr->sq_tids); free(merged_hdr->target_name); free(merged_hdr->target_len); free(merged_hdr); return NULL; } /* Some handy kstring manipulating functions */ // Append char range to kstring static inline int range_to_ks(const char *src, int from, int to, kstring_t *dest) { return kputsn(src + from, to - from, dest) != to - from; } // Append a header line match to kstring static inline int match_to_ks(const char *src, const hdr_match_t *match, kstring_t *dest) { return range_to_ks(src, match->rm_so, match->rm_eo, dest); } // Append a kstring to a kstring static inline int ks_to_ks(kstring_t *src, kstring_t *dest) { return kputsn(ks_str(src), ks_len(src), dest) != ks_len(src); } /* * Generate a unique ID by appending a random suffix to a given prefix. * existing_ids is the set of IDs that are already in use. * If always_add_suffix is true, the suffix will always be included. * If false, prefix will be returned unchanged if it isn't in existing_ids. */ static int gen_unique_id(char *prefix, khash_t(cset) *existing_ids, bool always_add_suffix, kstring_t *dest) { khiter_t iter; if (!always_add_suffix) { // Try prefix on its own first iter = kh_get(cset, existing_ids, prefix); if (iter == kh_end(existing_ids)) { // prefix isn't used yet dest->l = 0; if (kputs(prefix, dest) == EOF) return -1; return 0; } } do { dest->l = 0; ksprintf(dest, "%s-%0lX", prefix, lrand48()); iter = kh_get(cset, existing_ids, ks_str(dest)); } while (iter != kh_end(existing_ids)); return 0; } /* * Add the @HD line to the new header * In practice the @HD line will come from the first input header. */ static int trans_tbl_add_hd(merged_header_t* merged_hdr, bam_hdr_t *translate) { hdr_match_t match = {0, 0}; // TODO: handle case when @HD needs merging. if (merged_hdr->have_hd) return 0; if (hdr_line_match(translate->text, "@HD", NULL, &match) != 0) { return 0; } if (match_to_ks(translate->text, &match, &merged_hdr->out_hd)) goto memfail; if (kputc('\n', &merged_hdr->out_hd) == EOF) goto memfail; merged_hdr->have_hd = true; return 0; memfail: perror(__func__); return -1; } static inline int grow_target_list(merged_header_t* merged_hdr) { size_t new_size; char **new_names; uint32_t *new_len; new_size = merged_hdr->targets_sz * 2; new_names = realloc(merged_hdr->target_name, sizeof(*new_names) * new_size); if (!new_names) goto fail; merged_hdr->target_name = new_names; new_len = realloc(merged_hdr->target_len, sizeof(*new_len) * new_size); if (!new_len) goto fail; merged_hdr->target_len = new_len; merged_hdr->targets_sz = new_size; return 0; fail: perror(__func__); return -1; } /* * Add @SQ records to the translation table. * * Go through the target list for the input header. Any new targets found * are added to the output header target list. At the same time, a mapping * from the input to output target ids is stored in tbl. * * If any new targets are found, the header text is scanned to find the * corresponding @SQ records. They are then copied into the * merged_hdr->out_text kstring (which will eventually become the * output header text). * * Returns 0 on success, -1 on failure. */ static int trans_tbl_add_sq(merged_header_t* merged_hdr, bam_hdr_t *translate, trans_tbl_t* tbl) { kstring_t *out_text = &merged_hdr->out_sq; khash_t(c2i)* sq_tids = merged_hdr->sq_tids; hdr_match_t *new_sq_matches = NULL; char *text; hdr_match_t matches[2]; int32_t i, missing; int32_t old_n_targets = merged_hdr->n_targets; khiter_t iter; int min_tid = -1; // Fill in the tid part of the translation table, adding new targets // to the merged header as we go. for (i = 0; i < translate->n_targets; ++i) { // Check if it's a new target. iter = kh_get(c2i, sq_tids, translate->target_name[i]); if (iter == kh_end(sq_tids)) { int ret; // Append missing entries to out_hdr if (merged_hdr->n_targets == merged_hdr->targets_sz) { if (grow_target_list(merged_hdr)) goto fail; } merged_hdr->target_name[merged_hdr->n_targets] = strdup(translate->target_name[i]); if (merged_hdr->target_name[merged_hdr->n_targets] == NULL) goto memfail; merged_hdr->target_len[merged_hdr->n_targets] = translate->target_len[i]; // Record the new identifier for reference below, // and when building the ttable for other inputs. iter = kh_put(c2i, sq_tids, merged_hdr->target_name[merged_hdr->n_targets], &ret); if (ret < 0) { free(merged_hdr->target_name[merged_hdr->n_targets]); goto memfail; } assert(ret > 0); // Should not be in hash already. kh_value(sq_tids, iter) = merged_hdr->n_targets; tbl->tid_trans[i] = merged_hdr->n_targets++; } else { tbl->tid_trans[i] = kh_value(sq_tids, iter); } if (tbl->tid_trans[i] > min_tid) { min_tid = tbl->tid_trans[i]; } else { tbl->lost_coord_sort = true; } } if (merged_hdr->n_targets == old_n_targets) return 0; // Everything done if no new targets. // Otherwise, find @SQ lines in translate->text for all newly added targets. new_sq_matches = malloc((merged_hdr->n_targets - old_n_targets) * sizeof(*new_sq_matches)); if (new_sq_matches == NULL) goto memfail; for (i = 0; i < merged_hdr->n_targets - old_n_targets; i++) { new_sq_matches[i].rm_so = new_sq_matches[i].rm_eo = -1; } text = translate->text; while (hdr_line_match(text, "@SQ", "SN", matches) == 0) { // matches[0] is whole line, matches[1] is SN value. // This is a bit disgusting, but avoids a copy... char c = text[matches[1].rm_eo]; int idx; text[matches[1].rm_eo] = '\0'; // Look up the SN value in the sq_tids hash. iter = kh_get(c2i, sq_tids, text + matches[1].rm_so); text[matches[1].rm_eo] = c; // restore text if (iter == kh_end(sq_tids)) { // Warn about this, but it's not really fatal. fprintf(stderr, "[W::%s] @SQ SN (%.*s) found in text header but not binary header.\n", __func__, (int) (matches[1].rm_eo - matches[1].rm_so), text + matches[1].rm_so); text += matches[0].rm_eo; continue; // Skip to next } idx = kh_value(sq_tids, iter); if (idx >= old_n_targets) { // is a new SQ, so record position so we can add it to out_text. assert(idx < merged_hdr->n_targets); ptrdiff_t off = text - translate->text; new_sq_matches[idx - old_n_targets].rm_so = matches[0].rm_so + off; new_sq_matches[idx - old_n_targets].rm_eo = matches[0].rm_eo + off; } // Carry on searching from end of current match text += matches[0].rm_eo; } // Check if any new targets have been missed missing = 0; for (i = 0; i < merged_hdr->n_targets - old_n_targets; i++) { if (new_sq_matches[i].rm_so >= 0) { if (match_to_ks(translate->text, &new_sq_matches[i], out_text)) goto memfail; if (kputc('\n', out_text) == EOF) goto memfail; } else { fprintf(stderr, "[E::%s] @SQ SN (%s) found in binary header but not text header.\n", __func__, merged_hdr->target_name[i + old_n_targets]); missing++; } } if (missing) goto fail; free(new_sq_matches); return 0; memfail: perror(__func__); fail: free(new_sq_matches); return -1; } /* * Common code for setting up RG and PG record ID tag translation. * * is_rg is true for RG translation, false for PG. * translate is the input bam header * merge is true if tags with the same ID are to be merged. * known_ids is the set of IDs already in the output header. * id_map is the translation map from input header IDs to output header IDs * If override is set, it will be used to replace the existing ID (RG only) * * known_ids and id_map have entries for the new IDs added to them. * * Return value is a linked list of header lines with the translated IDs, * or NULL if something went wrong (probably out of memory). * */ static klist_t(hdrln) * trans_rg_pg(bool is_rg, bam_hdr_t *translate, bool merge, khash_t(cset)* known_ids, khash_t(c2c)* id_map, char *override) { hdr_match_t matches[2]; khiter_t iter; const char *text = translate->text; const char *rec_type = is_rg ? "@RG" : "@PG"; klist_t(hdrln) *hdr_lines; hdr_lines = kl_init(hdrln); // Search through translate's header while (hdr_line_match(text, rec_type, "ID", matches) == 0) { // matches[0] is the whole @RG/PG line; matches[1] is the ID field value kstring_t orig_id = { 0, 0, NULL }; // ID in original header kstring_t transformed_id = { 0, 0, NULL }; // ID in output header char *map_value; // Value to store in id_map bool id_changed; // Have we changed the ID? bool not_found_in_output; // ID isn't in the output header (yet) // Take a copy of the ID as we'll need it for a hash key. if (match_to_ks(text, &matches[1], &orig_id)) goto memfail; // is our matched ID in our output ID set already? iter = kh_get(cset, known_ids, ks_str(&orig_id)); not_found_in_output = (iter == kh_end(known_ids)); if (override) { // Override original ID (RG only) #ifdef OVERRIDE_DOES_NOT_MERGE if (gen_unique_id(override, known_ids, false, &transformed_id)) goto memfail; not_found_in_output = true; // As ID now unique #else if (kputs(override, &transformed_id) == EOF) goto memfail; // Know about override already? iter = kh_get(cset, known_ids, ks_str(&transformed_id)); not_found_in_output = (iter == kh_end(known_ids)); #endif id_changed = true; } else { if ( not_found_in_output || merge) { // Not in there or merging so can add it as 1-1 mapping if (ks_to_ks(&orig_id, &transformed_id)) goto memfail; id_changed = false; } else { // It's in there so we need to transform it by appending // a random number to the id if (gen_unique_id(ks_str(&orig_id), known_ids, true, &transformed_id)) goto memfail; id_changed = true; not_found_in_output = true; // As ID now unique } } // Does this line need to go into our output header? if (not_found_in_output) { // Take matched line and replace ID with transformed_id kstring_t new_hdr_line = { 0, 0, NULL }; if (!id_changed) { // Can just copy if (match_to_ks(text, &matches[0], &new_hdr_line)) goto memfail; } else { // Substitute new name for original if (range_to_ks(text, matches[0].rm_so, matches[1].rm_so, &new_hdr_line)) goto memfail; if (ks_to_ks(&transformed_id, &new_hdr_line)) goto memfail; if (range_to_ks(text, matches[1].rm_eo, matches[0].rm_eo, &new_hdr_line)) goto memfail; } // append line to output linked list char** ln = kl_pushp(hdrln, hdr_lines); *ln = ks_release(&new_hdr_line); // Give away to linked list // Need to add it to known_ids set int in_there = 0; iter = kh_put(cset, known_ids, ks_str(&transformed_id), &in_there); if (in_there < 0) goto memfail; assert(in_there > 0); // Should not already be in the map map_value = ks_release(&transformed_id); } else { // Use existing string in id_map assert(kh_exist(known_ids, iter)); map_value = kh_key(known_ids, iter); free(ks_release(&transformed_id)); } // Insert it into our translation map int in_there = 0; iter = kh_put(c2c, id_map, ks_release(&orig_id), &in_there); kh_value(id_map, iter) = map_value; text += matches[0].rm_eo; // next! } // If there are no RG lines in the file and we are overriding add one if (is_rg && override && kl_begin(hdr_lines) == NULL) { kstring_t new_id = {0, 0, NULL}; kstring_t line = {0, 0, NULL}; kstring_t empty = {0, 0, NULL}; int in_there = 0; char** ln; // Get the new ID if (gen_unique_id(override, known_ids, false, &new_id)) goto memfail; // Make into a header line and add to linked list ksprintf(&line, "@RG\tID:%s", ks_str(&new_id)); ln = kl_pushp(hdrln, hdr_lines); *ln = ks_release(&line); // Put into known_ids set iter = kh_put(cset, known_ids, ks_str(&new_id), &in_there); if (in_there < 0) goto memfail; assert(in_there > 0); // Should be a new entry // Put into translation map (key is empty string) if (kputs("", &empty) == EOF) goto memfail; iter = kh_put(c2c, id_map, ks_release(&empty), &in_there); if (in_there < 0) goto memfail; assert(in_there > 0); // Should be a new entry kh_value(id_map, iter) = ks_release(&new_id); } return hdr_lines; memfail: perror(__func__); if (hdr_lines) kl_destroy(hdrln, hdr_lines); return NULL; } /* * Common code for completing RG and PG record translation. * * Input is a list of header lines, and the mapping from input to * output @PG record IDs. * * RG and PG records can contain tags that cross-reference to other @PG * records. This fixes the tags to contain the new IDs before adding * them to the output header text. */ static int finish_rg_pg(bool is_rg, klist_t(hdrln) *hdr_lines, khash_t(c2c)* pg_map, kstring_t *out_text) { const char *search = is_rg ? "\tPG:" : "\tPP:"; khiter_t idx; char *line = NULL; while ((kl_shift(hdrln, hdr_lines, &line)) == 0) { char *id = strstr(line, search); // Look for tag to fix int pos1 = 0, pos2 = 0; char *new_id = NULL; if (id) { // Found a tag. Look up the value in the translation map // to see what it should be changed to in the output file. char *end, tmp; id += 4; // Point to value end = strchr(id, '\t'); // Find end of tag if (!end) end = id + strlen(id); tmp = *end; *end = '\0'; // Temporarily get the value on its own. // Look-up in translation table idx = kh_get(c2c, pg_map, id); if (idx == kh_end(pg_map)) { // Not found, warn. fprintf(stderr, "[W::%s] Tag %s%s not found in @PG records\n", __func__, search + 1, id); } else { // Remember new id and splice points on original string new_id = kh_value(pg_map, idx); pos1 = id - line; pos2 = end - line; } *end = tmp; // Restore string } // Copy line to output: // line[0..pos1), new_id (if not NULL), line[pos2..end), '\n' if (pos1 && range_to_ks(line, 0, pos1, out_text)) goto memfail; if (new_id && kputs(new_id, out_text) == EOF) goto memfail; if (kputs(line + pos2, out_text) == EOF) goto memfail; if (kputc('\n', out_text) == EOF) goto memfail; free(line); // No longer needed line = NULL; } return 0; memfail: perror(__func__); free(line); // Prevent leakage as no longer on list return -1; } /* * Build the translation table for an input *am file. This stores mappings * which allow IDs to be converted from those used in the input file * to the ones which will be used in the output. The mappings are for: * Reference sequence IDs (for @SQ records) * @RG record ID tags * @PG record ID tags * * At the same time, new header text is built up by copying records * from the input bam file. This will eventually become the header for * the output file. When copied, the ID tags for @RG and @PG records * are replaced with their values. The @PG PP: and @RG PG: tags * are also modified if necessary. * * merged_hdr holds state on the output header (which IDs are present, etc.) * translate is the input header * tbl is the translation table that gets filled in. * merge_rg controls merging of @RG records * merge_pg controls merging of @PG records * If rg_override is not NULL, it will be used to replace the existing @RG ID * * Returns 0 on success, -1 on failure. */ static int trans_tbl_init(merged_header_t* merged_hdr, bam_hdr_t* translate, trans_tbl_t* tbl, bool merge_rg, bool merge_pg, char* rg_override) { klist_t(hdrln) *rg_list = NULL; klist_t(hdrln) *pg_list = NULL; tbl->n_targets = translate->n_targets; tbl->rg_trans = tbl->pg_trans = NULL; tbl->tid_trans = (int*)calloc(translate->n_targets, sizeof(int)); if (tbl->tid_trans == NULL) goto memfail; tbl->rg_trans = kh_init(c2c); if (tbl->rg_trans == NULL) goto memfail; tbl->pg_trans = kh_init(c2c); if (tbl->pg_trans == NULL) goto memfail; tbl->lost_coord_sort = false; // Get the @HD record (if not there already). if (trans_tbl_add_hd(merged_hdr, translate)) goto fail; // Fill in map and add header lines for @SQ records if (trans_tbl_add_sq(merged_hdr, translate, tbl)) goto fail; // Get translated header lines and fill in map for @RG records rg_list = trans_rg_pg(true, translate, merge_rg, merged_hdr->rg_ids, tbl->rg_trans, rg_override); if (!rg_list) goto fail; // Get translated header lines and fill in map for @PG records pg_list = trans_rg_pg(false, translate, merge_pg, merged_hdr->pg_ids, tbl->pg_trans, NULL); // Fix-up PG: tags in the new @RG records and add to output if (finish_rg_pg(true, rg_list, tbl->pg_trans, &merged_hdr->out_rg)) goto fail; // Fix-up PP: tags in the new @PG records and add to output if (finish_rg_pg(false, pg_list, tbl->pg_trans, &merged_hdr->out_pg)) goto fail; kl_destroy(hdrln, rg_list); rg_list = NULL; kl_destroy(hdrln, pg_list); pg_list = NULL; // Just append @CO headers without translation const char *line, *end_pointer; for (line = translate->text; *line; line = end_pointer + 1) { end_pointer = strchr(line, '\n'); if (strncmp(line, "@CO", 3) == 0) { if (end_pointer) { if (kputsn(line, end_pointer - line + 1, &merged_hdr->out_co) == EOF) goto memfail; } else { // Last line with no trailing '\n' if (kputs(line, &merged_hdr->out_co) == EOF) goto memfail; if (kputc('\n', &merged_hdr->out_co) == EOF) goto memfail; } } if (end_pointer == NULL) break; } return 0; memfail: perror(__func__); fail: trans_tbl_destroy(tbl); if (rg_list) kl_destroy(hdrln, rg_list); if (pg_list) kl_destroy(hdrln, pg_list); return -1; } static inline void move_kstr_to_text(char **text, kstring_t *ks) { memcpy(*text, ks_str(ks), ks_len(ks)); *text += ks_len(ks); **text = '\0'; free(ks_release(ks)); } /* * Populate a bam_hdr_t struct from data in a merged_header_t. */ static bam_hdr_t * finish_merged_header(merged_header_t *merged_hdr) { size_t txt_sz; char *text; bam_hdr_t *hdr; // Check output text size txt_sz = (ks_len(&merged_hdr->out_hd) + ks_len(&merged_hdr->out_sq) + ks_len(&merged_hdr->out_rg) + ks_len(&merged_hdr->out_pg) + ks_len(&merged_hdr->out_co)); if (txt_sz >= INT32_MAX) { fprintf(stderr, "[%s] Output header text too long\n", __func__); return NULL; } // Allocate new header hdr = bam_hdr_init(); if (hdr == NULL) goto memfail; // Transfer targets arrays to new header hdr->n_targets = merged_hdr->n_targets; if (hdr->n_targets > 0) { // Try to shrink targets arrays to correct size hdr->target_name = realloc(merged_hdr->target_name, hdr->n_targets * sizeof(char*)); if (!hdr->target_name) hdr->target_name = merged_hdr->target_name; hdr->target_len = realloc(merged_hdr->target_len, hdr->n_targets * sizeof(uint32_t)); if (!hdr->target_len) hdr->target_len = merged_hdr->target_len; // These have either been freed by realloc() or, in the unlikely // event that failed, have had their ownership transferred to hdr merged_hdr->target_name = NULL; merged_hdr->target_len = NULL; } else { hdr->target_name = NULL; hdr->target_len = NULL; } // Allocate text text = hdr->text = malloc(txt_sz + 1); if (!text) goto memfail; // Put header text in order @HD, @SQ, @RG, @PG, @CO move_kstr_to_text(&text, &merged_hdr->out_hd); move_kstr_to_text(&text, &merged_hdr->out_sq); move_kstr_to_text(&text, &merged_hdr->out_rg); move_kstr_to_text(&text, &merged_hdr->out_pg); move_kstr_to_text(&text, &merged_hdr->out_co); hdr->l_text = txt_sz; return hdr; memfail: perror(__func__); bam_hdr_destroy(hdr); return NULL; } /* * Free a merged_header_t struct and all associated data. * * Note that the keys to the rg_ids and pg_ids sets are also used as * values in the translation tables. This function should therefore not * be called until the translation tables are no longer needed. */ static void free_merged_header(merged_header_t *merged_hdr) { size_t i; khiter_t iter; if (!merged_hdr) return; free(ks_release(&merged_hdr->out_hd)); free(ks_release(&merged_hdr->out_sq)); free(ks_release(&merged_hdr->out_rg)); free(ks_release(&merged_hdr->out_pg)); free(ks_release(&merged_hdr->out_co)); if (merged_hdr->target_name) { for (i = 0; i < merged_hdr->n_targets; i++) { free(merged_hdr->target_name[i]); } free(merged_hdr->target_name); } free(merged_hdr->target_len); kh_destroy(c2i, merged_hdr->sq_tids); if (merged_hdr->rg_ids) { for (iter = kh_begin(merged_hdr->rg_ids); iter != kh_end(merged_hdr->rg_ids); ++iter) { if (kh_exist(merged_hdr->rg_ids, iter)) free(kh_key(merged_hdr->rg_ids, iter)); } kh_destroy(cset, merged_hdr->rg_ids); } if (merged_hdr->pg_ids) { for (iter = kh_begin(merged_hdr->pg_ids); iter != kh_end(merged_hdr->pg_ids); ++iter) { if (kh_exist(merged_hdr->pg_ids, iter)) free(kh_key(merged_hdr->pg_ids, iter)); } kh_destroy(cset, merged_hdr->pg_ids); } free(merged_hdr); } static void bam_translate(bam1_t* b, trans_tbl_t* tbl) { // Update target id if not unmapped tid if ( b->core.tid >= 0 ) { b->core.tid = tbl->tid_trans[b->core.tid]; } if ( b->core.mtid >= 0 ) { b->core.mtid = tbl->tid_trans[b->core.mtid]; } // If we have a RG update it uint8_t *rg = bam_aux_get(b, "RG"); if (rg) { char* decoded_rg = bam_aux2Z(rg); khiter_t k = kh_get(c2c, tbl->rg_trans, decoded_rg); if (k != kh_end(tbl->rg_trans)) { char* translate_rg = kh_value(tbl->rg_trans,k); bam_aux_del(b, rg); if (translate_rg) { bam_aux_append(b, "RG", 'Z', strlen(translate_rg) + 1, (uint8_t*)translate_rg); } } else { char *tmp = strdup(decoded_rg); fprintf(stderr, "[bam_translate] RG tag \"%s\" on read \"%s\" encountered " "with no corresponding entry in header, tag lost. " "Unknown tags are only reported once per input file for " "each tag ID.\n", decoded_rg, bam_get_qname(b)); bam_aux_del(b, rg); // Prevent future whinges if (tmp) { int in_there = 0; k = kh_put(c2c, tbl->rg_trans, tmp, &in_there); if (in_there > 0) kh_value(tbl->rg_trans, k) = NULL; } } } // If we have a PG update it uint8_t *pg = bam_aux_get(b, "PG"); if (pg) { char* decoded_pg = bam_aux2Z(pg); khiter_t k = kh_get(c2c, tbl->pg_trans, decoded_pg); if (k != kh_end(tbl->pg_trans)) { char* translate_pg = kh_value(tbl->pg_trans,k); bam_aux_del(b, pg); if (translate_pg) { bam_aux_append(b, "PG", 'Z', strlen(translate_pg) + 1, (uint8_t*)translate_pg); } } else { char *tmp = strdup(decoded_pg); fprintf(stderr, "[bam_translate] PG tag \"%s\" on read \"%s\" encountered " "with no corresponding entry in header, tag lost. " "Unknown tags are only reported once per input file for " "each tag ID.\n", decoded_pg, bam_get_qname(b)); bam_aux_del(b, pg); // Prevent future whinges if (tmp) { int in_there = 0; k = kh_put(c2c, tbl->pg_trans, tmp, &in_there); if (in_there > 0) kh_value(tbl->pg_trans, k) = NULL; } } } } int* rtrans_build(int n, int n_targets, trans_tbl_t* translation_tbl) { // Create reverse translation table for tids int* rtrans = (int*)malloc(sizeof(int32_t)*n*n_targets); const int32_t NOTID = INT32_MIN; memset_pattern4((void*)rtrans, &NOTID, sizeof(int32_t)*n*n_targets); int i; for (i = 0; i < n; ++i) { int j; for (j = 0; j < (translation_tbl+i)->n_targets; ++j) { if ((translation_tbl+i)->tid_trans[j] != -1) { rtrans[i*n_targets + (translation_tbl+i)->tid_trans[j]] = j; } } } return rtrans; } #define MERGE_RG 1 // Attach RG tag based on filename #define MERGE_UNCOMP 2 // Generate uncompressed BAM #define MERGE_LEVEL1 4 // Compress the BAM at level 1 (fast) mode #define MERGE_FORCE 8 // Overwrite output BAM if it exists #define MERGE_COMBINE_RG 16 // Combine RG tags frather than redefining them #define MERGE_COMBINE_PG 32 // Combine PG tags frather than redefining them /* * How merging is handled * * If a hheader is defined use we will use that as our output header * otherwise we use the first header from the first input file. * * Now go through each file and create a translation table for that file for: * -RG * -tid * -PG tags * * Then whenever we read a record from a bam we translate that read before * stashing it in the hash. * * In the actual merge, a read is read from each input file, translated and * stashed in the hash. This assumes that all input files are sorted in the * same way. Next we just extract the next position ordered read from the * hash, and replace it if there are still reads left in it's source input * file. Finally we write our chosen read it to the output file. */ /*! @abstract Merge multiple sorted BAM. @param is_by_qname whether to sort by query name @param out output BAM file name @param mode sam_open() mode to be used to create the final output file (overrides level settings from UNCOMP and LEVEL1 flags) @param headers name of SAM file from which to copy '@' header lines, or NULL to copy them from the first file to be merged @param n number of files to be merged @param fn names of files to be merged @param flag flags that control how the merge is undertaken @param reg region to merge @param n_threads number of threads to use (passed to htslib) @param in_fmt format options for input files @param out_fmt output file format and options @discussion Padding information may NOT correctly maintained. This function is NOT thread safe. */ int bam_merge_core2(int by_qname, const char *out, const char *mode, const char *headers, int n, char * const *fn, int flag, const char *reg, int n_threads, const htsFormat *in_fmt, const htsFormat *out_fmt) { samFile *fpout, **fp; heap1_t *heap; bam_hdr_t *hout = NULL; bam_hdr_t *hin = NULL; int i, j, *RG_len = NULL; uint64_t idx = 0; char **RG = NULL; hts_itr_t **iter = NULL; bam_hdr_t **hdr = NULL; trans_tbl_t *translation_tbl = NULL; merged_header_t *merged_hdr = init_merged_header(); if (!merged_hdr) return -1; // Is there a specified pre-prepared header to use for output? if (headers) { samFile* fpheaders = sam_open(headers, "r"); if (fpheaders == NULL) { const char *message = strerror(errno); fprintf(stderr, "[bam_merge_core] cannot open '%s': %s\n", headers, message); return -1; } hin = sam_hdr_read(fpheaders); sam_close(fpheaders); if (hin == NULL) { fprintf(stderr, "[bam_merge_core] couldn't read headers for '%s'\n", headers); return -1; } } g_is_by_qname = by_qname; fp = (samFile**)calloc(n, sizeof(samFile*)); heap = (heap1_t*)calloc(n, sizeof(heap1_t)); iter = (hts_itr_t**)calloc(n, sizeof(hts_itr_t*)); hdr = (bam_hdr_t**)calloc(n, sizeof(bam_hdr_t*)); translation_tbl = (trans_tbl_t*)calloc(n, sizeof(trans_tbl_t)); RG = (char**)calloc(n, sizeof(char*)); // prepare RG tag from file names if (flag & MERGE_RG) { RG_len = (int*)calloc(n, sizeof(int)); for (i = 0; i != n; ++i) { int l = strlen(fn[i]); const char *s = fn[i]; if (l > 4 && (strcmp(s + l - 4, ".bam") == 0 || strcmp(s + l - 4, ".sam") == 0)) l -= 4; if (l > 5 && strcmp(s + l - 5, ".cram") == 0) l -= 5; for (j = l - 1; j >= 0; --j) if (s[j] == '/') break; ++j; l -= j; RG[i] = (char*)calloc(l + 1, 1); RG_len[i] = l; strncpy(RG[i], s + j, l); } } if (hin) { // Popluate merged_hdr from the pre-prepared header trans_tbl_t dummy; int res; res = trans_tbl_init(merged_hdr, hin, &dummy, flag & MERGE_COMBINE_RG, flag & MERGE_COMBINE_PG, NULL); trans_tbl_destroy(&dummy); if (res) return -1; // FIXME: memory leak } // open and read the header from each file for (i = 0; i < n; ++i) { bam_hdr_t *hin; fp[i] = sam_open_format(fn[i], "r", in_fmt); if (fp[i] == NULL) { int j; fprintf(stderr, "[bam_merge_core] fail to open file %s\n", fn[i]); for (j = 0; j < i; ++j) { bam_hdr_destroy(hdr[i]); sam_close(fp[j]); } free(fp); free(heap); // FIXME: possible memory leak return -1; } hin = sam_hdr_read(fp[i]); if (hin == NULL) { fprintf(stderr, "[bam_merge_core] failed to read header for '%s'\n", fn[i]); for (j = 0; j < i; ++j) { bam_hdr_destroy(hdr[i]); sam_close(fp[j]); } free(fp); free(heap); // FIXME: possible memory leak return -1; } if (trans_tbl_init(merged_hdr, hin, translation_tbl+i, flag & MERGE_COMBINE_RG, flag & MERGE_COMBINE_PG, RG[i])) return -1; // FIXME: memory leak // TODO sam_itr_next() doesn't yet work for SAM files, // so for those keep the headers around for use with sam_read1() if (hts_get_format(fp[i])->format == sam) hdr[i] = hin; else { bam_hdr_destroy(hin); hdr[i] = NULL; } if ((translation_tbl+i)->lost_coord_sort && !by_qname) { fprintf(stderr, "[bam_merge_core] Order of targets in file %s caused coordinate sort to be lost\n", fn[i]); } } // Did we get an @HD line? if (!merged_hdr->have_hd) { fprintf(stderr, "[W::%s] No @HD tag found.\n", __func__); /* FIXME: Should we add an @HD line here, and if so what should we put in it? Ideally we want a way of getting htslib to tell us the SAM version number to assume given no @HD line. Is it also safe to assume that the output is coordinate sorted? SO: is optional so we don't have to have it.*/ /* ksprintf(&merged_hdr->out_hd, "@HD\tVN:1.5\tSO:coordinate\n"); */ } // Transform the header into standard form hout = finish_merged_header(merged_hdr); if (!hout) return -1; // FIXME: memory leak // If we're only merging a specified region move our iters to start at that point if (reg) { int* rtrans = rtrans_build(n, hout->n_targets, translation_tbl); int tid, beg, end; const char *name_lim = hts_parse_reg(reg, &beg, &end); if (name_lim) { char *name = malloc(name_lim - reg + 1); memcpy(name, reg, name_lim - reg); name[name_lim - reg] = '\0'; tid = bam_name2id(hout, name); free(name); } else { // not parsable as a region, but possibly a sequence named "foo:a" tid = bam_name2id(hout, reg); beg = 0; end = INT_MAX; } if (tid < 0) { if (name_lim) fprintf(stderr, "[%s] Region \"%s\" specifies an unknown reference name\n", __func__, reg); else fprintf(stderr, "[%s] Badly formatted region: \"%s\"\n", __func__, reg); return -1; } for (i = 0; i < n; ++i) { hts_idx_t *idx = sam_index_load(fp[i], fn[i]); // (rtrans[i*n+tid]) Look up what hout tid translates to in input tid space int mapped_tid = rtrans[i*hout->n_targets+tid]; if (idx == NULL) { fprintf(stderr, "[%s] failed to load index for %s. Random alignment retrieval only works for indexed BAM or CRAM files.\n", __func__, fn[i]); return -1; } if (mapped_tid != INT32_MIN) { iter[i] = sam_itr_queryi(idx, mapped_tid, beg, end); } else { iter[i] = sam_itr_queryi(idx, HTS_IDX_NONE, 0, 0); } hts_idx_destroy(idx); if (iter[i] == NULL) break; } free(rtrans); } else { for (i = 0; i < n; ++i) { if (hdr[i] == NULL) { iter[i] = sam_itr_queryi(NULL, HTS_IDX_REST, 0, 0); if (iter[i] == NULL) break; } else iter[i] = NULL; } } if (i < n) { fprintf(stderr, "[%s] Memory allocation failed\n", __func__); return -1; } // Load the first read from each file into the heap for (i = 0; i < n; ++i) { heap1_t *h = heap + i; h->i = i; h->b = bam_init1(); if ((iter[i]? sam_itr_next(fp[i], iter[i], h->b) : sam_read1(fp[i], hdr[i], h->b)) >= 0) { bam_translate(h->b, translation_tbl + i); h->pos = ((uint64_t)h->b->core.tid<<32) | (uint32_t)((int32_t)h->b->core.pos+1)<<1 | bam_is_rev(h->b); h->idx = idx++; } else { h->pos = HEAP_EMPTY; bam_destroy1(h->b); h->b = NULL; } } // Open output file and write header if ((fpout = sam_open_format(out, mode, out_fmt)) == 0) { fprintf(stderr, "[%s] fail to create the output file.\n", __func__); return -1; } sam_hdr_write(fpout, hout); if (!(flag & MERGE_UNCOMP)) hts_set_threads(fpout, n_threads); // Begin the actual merge ks_heapmake(heap, n, heap); while (heap->pos != HEAP_EMPTY) { bam1_t *b = heap->b; if (flag & MERGE_RG) { uint8_t *rg = bam_aux_get(b, "RG"); if (rg) bam_aux_del(b, rg); bam_aux_append(b, "RG", 'Z', RG_len[heap->i] + 1, (uint8_t*)RG[heap->i]); } sam_write1(fpout, hout, b); if ((j = (iter[heap->i]? sam_itr_next(fp[heap->i], iter[heap->i], b) : sam_read1(fp[heap->i], hdr[heap->i], b))) >= 0) { bam_translate(b, translation_tbl + heap->i); heap->pos = ((uint64_t)b->core.tid<<32) | (uint32_t)((int)b->core.pos+1)<<1 | bam_is_rev(b); heap->idx = idx++; } else if (j == -1) { heap->pos = HEAP_EMPTY; bam_destroy1(heap->b); heap->b = NULL; } else fprintf(stderr, "[bam_merge_core] '%s' is truncated. Continue anyway.\n", fn[heap->i]); ks_heapadjust(heap, 0, n, heap); } // Clean up and close if (flag & MERGE_RG) { for (i = 0; i != n; ++i) free(RG[i]); free(RG_len); } for (i = 0; i < n; ++i) { trans_tbl_destroy(translation_tbl + i); hts_itr_destroy(iter[i]); bam_hdr_destroy(hdr[i]); sam_close(fp[i]); } bam_hdr_destroy(hin); bam_hdr_destroy(hout); free_merged_header(merged_hdr); sam_close(fpout); free(RG); free(translation_tbl); free(fp); free(heap); free(iter); free(hdr); return 0; } // Unused here but may be used by legacy samtools-using third-party code int bam_merge_core(int by_qname, const char *out, const char *headers, int n, char * const *fn, int flag, const char *reg) { char mode[12]; strcpy(mode, "wb"); if (flag & MERGE_UNCOMP) strcat(mode, "0"); else if (flag & MERGE_LEVEL1) strcat(mode, "1"); return bam_merge_core2(by_qname, out, mode, headers, n, fn, flag, reg, 0, NULL, NULL); } static void merge_usage(FILE *to) { fprintf(to, "Usage: samtools merge [-nurlf] [-h inh.sam] [-b ] [ ... ]\n" "\n" "Options:\n" " -n Sort by read names\n" " -r Attach RG tag (inferred from file names)\n" " -u Uncompressed BAM output\n" " -f Overwrite the output BAM if exist\n" " -1 Compress level 1\n" " -l INT Compression level, from 0 to 9 [-1]\n" " -R STR Merge file in the specified region STR [all]\n" " -h FILE Copy the header in FILE to [in1.bam]\n" " -c Combine @RG headers with colliding IDs [alter IDs to be distinct]\n" " -p Combine @PG headers with colliding IDs [alter IDs to be distinct]\n" " -s VALUE Override random seed\n" " -b FILE List of input BAM filenames, one per line [null]\n" " -@, --threads INT\n" " Number of BAM/CRAM compression threads [0]\n"); sam_global_opt_help(to, "-.O.."); } int bam_merge(int argc, char *argv[]) { int c, is_by_qname = 0, flag = 0, ret = 0, n_threads = 0, level = -1; char *fn_headers = NULL, *reg = NULL, mode[12]; long random_seed = (long)time(NULL); char** fn = NULL; int fn_size = 0; sam_global_args ga = SAM_GLOBAL_ARGS_INIT; static const struct option lopts[] = { SAM_OPT_GLOBAL_OPTIONS('-', 0, 'O', 0, 0), { "threads", required_argument, NULL, '@' }, { NULL, 0, NULL, 0 } }; if (argc == 1) { merge_usage(stdout); return 0; } while ((c = getopt_long(argc, argv, "h:nru1R:f@:l:cps:b:O:", lopts, NULL)) >= 0) { switch (c) { case 'r': flag |= MERGE_RG; break; case 'f': flag |= MERGE_FORCE; break; case 'h': fn_headers = strdup(optarg); break; case 'n': is_by_qname = 1; break; case '1': flag |= MERGE_LEVEL1; level = 1; break; case 'u': flag |= MERGE_UNCOMP; level = 0; break; case 'R': reg = strdup(optarg); break; case 'l': level = atoi(optarg); break; case '@': n_threads = atoi(optarg); break; case 'c': flag |= MERGE_COMBINE_RG; break; case 'p': flag |= MERGE_COMBINE_PG; break; case 's': random_seed = atol(optarg); break; case 'b': { // load the list of files to read int nfiles; char **fn_read = hts_readlines(optarg, &nfiles); if (fn_read) { // Append to end of array fn = realloc(fn, (fn_size+nfiles) * sizeof(char*)); if (fn == NULL) { ret = 1; goto end; } memcpy(fn+fn_size, fn_read, nfiles * sizeof(char*)); fn_size += nfiles; } else { fprintf(stderr, "[%s] Invalid file list \"%s\"\n", __func__, optarg); ret = 1; } break; } default: if (parse_sam_global_opt(c, optarg, lopts, &ga) == 0) break; /* else fall-through */ case '?': merge_usage(stderr); return 1; } } if ( argc - optind < 1 ) { fprintf(stderr, "You must at least specify the output file.\n"); merge_usage(stderr); return 1; } srand48(random_seed); if (!(flag & MERGE_FORCE) && strcmp(argv[optind], "-")) { FILE *fp = fopen(argv[optind], "rb"); if (fp != NULL) { fclose(fp); fprintf(stderr, "[%s] File '%s' exists. Please apply '-f' to overwrite. Abort.\n", __func__, argv[optind]); return 1; } } int nargcfiles = argc - (optind+1); if (nargcfiles > 0) { // Add argc files to end of array fn = realloc(fn, (fn_size+nargcfiles) * sizeof(char*)); if (fn == NULL) { ret = 1; goto end; } memcpy(fn+fn_size, argv + (optind+1), nargcfiles * sizeof(char*)); } if (fn_size+nargcfiles < 1) { fprintf(stderr, "You must specify at least one (and usually two or more) input files.\n"); merge_usage(stderr); return 1; } strcpy(mode, "wb"); sam_open_mode(mode+1, argv[optind], NULL); if (level >= 0) sprintf(strchr(mode, '\0'), "%d", level < 9? level : 9); if (bam_merge_core2(is_by_qname, argv[optind], mode, fn_headers, fn_size+nargcfiles, fn, flag, reg, n_threads, &ga.in, &ga.out) < 0) ret = 1; end: if (fn_size > 0) { int i; for (i=0; i typedef bam1_t *bam1_p; static int change_SO(bam_hdr_t *h, const char *so) { char *p, *q, *beg = NULL, *end = NULL, *newtext; if (h->l_text > 3) { if (strncmp(h->text, "@HD", 3) == 0) { if ((p = strchr(h->text, '\n')) == 0) return -1; *p = '\0'; if ((q = strstr(h->text, "\tSO:")) != 0) { *p = '\n'; // change back if (strncmp(q + 4, so, p - q - 4) != 0) { beg = q; for (q += 4; *q != '\n' && *q != '\t'; ++q); end = q; } else return 0; // no need to change } else beg = end = p, *p = '\n'; } } if (beg == NULL) { // no @HD h->l_text += strlen(so) + 15; newtext = (char*)malloc(h->l_text + 1); sprintf(newtext, "@HD\tVN:1.3\tSO:%s\n", so); strcat(newtext, h->text); } else { // has @HD but different or no SO h->l_text = (beg - h->text) + (4 + strlen(so)) + (h->text + h->l_text - end); newtext = (char*)malloc(h->l_text + 1); strncpy(newtext, h->text, beg - h->text); sprintf(newtext + (beg - h->text), "\tSO:%s", so); strcat(newtext, end); } free(h->text); h->text = newtext; return 0; } // Function to compare reads and determine which one is < the other static inline int bam1_lt(const bam1_p a, const bam1_p b) { if (g_is_by_qname) { int t = strnum_cmp(bam_get_qname(a), bam_get_qname(b)); //return (t < 0 || (t == 0 && (a->core.flag&0xc0) < (b->core.flag&0xc0))); // Modified for RSEM by Bo Li return t < 0; } else return (((uint64_t)a->core.tid<<32|(a->core.pos+1)<<1|bam_is_rev(a)) < ((uint64_t)b->core.tid<<32|(b->core.pos+1)<<1|bam_is_rev(b))); } KSORT_INIT(sort, bam1_p, bam1_lt) typedef struct { size_t buf_len; const char *prefix; bam1_p *buf; const bam_hdr_t *h; int index; } worker_t; static void write_buffer(const char *fn, const char *mode, size_t l, bam1_p *buf, const bam_hdr_t *h, int n_threads, const htsFormat *fmt) { size_t i; samFile* fp; fp = sam_open_format(fn, mode, fmt); if (fp == NULL) return; sam_hdr_write(fp, h); if (n_threads > 1) hts_set_threads(fp, n_threads); for (i = 0; i < l; ++i) sam_write1(fp, h, buf[i]); sam_close(fp); } static void *worker(void *data) { worker_t *w = (worker_t*)data; char *name; ks_mergesort(sort, w->buf_len, w->buf, 0); name = (char*)calloc(strlen(w->prefix) + 20, 1); sprintf(name, "%s.%.4d.bam", w->prefix, w->index); write_buffer(name, "wb1", w->buf_len, w->buf, w->h, 0, NULL); // Consider using CRAM temporary files if the final output is CRAM. // Typically it is comparable speed while being smaller. // hts_opt opt[2] = { // {"version=3.0", CRAM_OPT_VERSION, {"3.0"}, NULL}, // {"no_ref", CRAM_OPT_NO_REF, {1}, NULL} // }; // opt[0].next = &opt[1]; // write_buffer(name, "wc1", w->buf_len, w->buf, w->h, 0, opt); free(name); return 0; } static int sort_blocks(int n_files, size_t k, bam1_p *buf, const char *prefix, const bam_hdr_t *h, int n_threads) { int i; size_t rest; bam1_p *b; pthread_t *tid; pthread_attr_t attr; worker_t *w; if (n_threads < 1) n_threads = 1; if (k < n_threads * 64) n_threads = 1; // use a single thread if we only sort a small batch of records pthread_attr_init(&attr); pthread_attr_setdetachstate(&attr, PTHREAD_CREATE_JOINABLE); w = (worker_t*)calloc(n_threads, sizeof(worker_t)); tid = (pthread_t*)calloc(n_threads, sizeof(pthread_t)); b = buf; rest = k; for (i = 0; i < n_threads; ++i) { w[i].buf_len = rest / (n_threads - i); w[i].buf = b; w[i].prefix = prefix; w[i].h = h; w[i].index = n_files + i; b += w[i].buf_len; rest -= w[i].buf_len; pthread_create(&tid[i], &attr, worker, &w[i]); } for (i = 0; i < n_threads; ++i) pthread_join(tid[i], 0); free(tid); free(w); return n_files + n_threads; } /*! @abstract Sort an unsorted BAM file based on the chromosome order and the leftmost position of an alignment @param is_by_qname whether to sort by query name @param fn name of the file to be sorted @param prefix prefix of the temporary files (prefix.NNNN.bam are written) @param fnout name of the final output file to be written @param modeout sam_open() mode to be used to create the final output file @param max_mem approxiate maximum memory (very inaccurate) @param in_fmt input file format options @param out_fmt output file format and options @return 0 for successful sorting, negative on errors @discussion It may create multiple temporary subalignment files and then merge them by calling bam_merge_core2(). This function is NOT thread safe. */ int bam_sort_core_ext(int is_by_qname, const char *fn, const char *prefix, const char *fnout, const char *modeout, size_t _max_mem, int n_threads, const htsFormat *in_fmt, const htsFormat *out_fmt) { int ret = -1, i, n_files = 0; size_t mem, max_k, k, max_mem; bam_hdr_t *header = NULL; samFile *fp; bam1_t *b, **buf; if (n_threads < 2) n_threads = 1; g_is_by_qname = is_by_qname; max_k = k = 0; mem = 0; max_mem = _max_mem * n_threads; buf = NULL; fp = sam_open_format(fn, "r", in_fmt); if (fp == NULL) { const char *message = strerror(errno); fprintf(stderr, "[bam_sort_core] fail to open '%s': %s\n", fn, message); return -2; } header = sam_hdr_read(fp); if (header == NULL) { fprintf(stderr, "[bam_sort_core] failed to read header for '%s'\n", fn); goto err; } if (is_by_qname) change_SO(header, "queryname"); else change_SO(header, "coordinate"); // write sub files for (;;) { if (k == max_k) { size_t kk, old_max = max_k; max_k = max_k? max_k<<1 : 0x10000; buf = (bam1_t**)realloc(buf, max_k * sizeof(bam1_t*)); for (kk = old_max; kk < max_k; ++kk) buf[kk] = NULL; } if (buf[k] == NULL) buf[k] = bam_init1(); b = buf[k]; if ((ret = sam_read1(fp, header, b)) < 0) break; if (b->l_data < b->m_data>>2) { // shrink b->m_data = b->l_data; kroundup32(b->m_data); b->data = (uint8_t*)realloc(b->data, b->m_data); } mem += sizeof(bam1_t) + b->m_data + sizeof(void*) + sizeof(void*); // two sizeof(void*) for the data allocated to pointer arrays ++k; if (mem >= max_mem) { n_files = sort_blocks(n_files, k, buf, prefix, header, n_threads); mem = k = 0; } } if (ret != -1) { fprintf(stderr, "[bam_sort_core] truncated file. Aborting.\n"); ret = -1; goto err; } // write the final output if (n_files == 0) { // a single block ks_mergesort(sort, k, buf, 0); write_buffer(fnout, modeout, k, buf, header, n_threads, out_fmt); } else { // then merge char **fns; n_files = sort_blocks(n_files, k, buf, prefix, header, n_threads); fprintf(stderr, "[bam_sort_core] merging from %d files...\n", n_files); fns = (char**)calloc(n_files, sizeof(char*)); for (i = 0; i < n_files; ++i) { fns[i] = (char*)calloc(strlen(prefix) + 20, 1); sprintf(fns[i], "%s.%.4d.bam", prefix, i); } if (bam_merge_core2(is_by_qname, fnout, modeout, NULL, n_files, fns, MERGE_COMBINE_RG|MERGE_COMBINE_PG, NULL, n_threads, in_fmt, out_fmt) < 0) { // Propagate bam_merge_core2() failure; it has already emitted a // message explaining the failure, so no further message is needed. goto err; } for (i = 0; i < n_files; ++i) { unlink(fns[i]); free(fns[i]); } free(fns); } ret = 0; err: // free for (k = 0; k < max_k; ++k) bam_destroy1(buf[k]); free(buf); bam_hdr_destroy(header); sam_close(fp); return ret; } // Unused here but may be used by legacy samtools-using third-party code int bam_sort_core(int is_by_qname, const char *fn, const char *prefix, size_t max_mem) { int ret; char *fnout = calloc(strlen(prefix) + 4 + 1, 1); sprintf(fnout, "%s.bam", prefix); ret = bam_sort_core_ext(is_by_qname, fn, prefix, fnout, "wb", max_mem, 0, NULL, NULL); free(fnout); return ret; } static void sort_usage(FILE *fp) { fprintf(fp, "Usage: samtools sort [options...] [in.bam]\n" "Options:\n" " -l INT Set compression level, from 0 (uncompressed) to 9 (best)\n" " -m INT Set maximum memory per thread; suffix K/M/G recognized [768M]\n" " -n Sort by read name\n" " -o FILE Write final output to FILE rather than standard output\n" " -T PREFIX Write temporary files to PREFIX.nnnn.bam\n" " -@, --threads INT\n" " Set number of sorting and compression threads [1]\n"); sam_global_opt_help(fp, "-.O.."); } int bam_sort(int argc, char *argv[]) { size_t max_mem = 768<<20; // 512MB int c, nargs, is_by_qname = 0, ret, o_seen = 0, n_threads = 0, level = -1; char *fnout = "-", modeout[12]; kstring_t tmpprefix = { 0, 0, NULL }; sam_global_args ga = SAM_GLOBAL_ARGS_INIT; static const struct option lopts[] = { SAM_OPT_GLOBAL_OPTIONS('-', 0, 'O', 0, 0), { "threads", required_argument, NULL, '@' }, { NULL, 0, NULL, 0 } }; while ((c = getopt_long(argc, argv, "l:m:no:O:T:@:", lopts, NULL)) >= 0) { switch (c) { case 'o': fnout = optarg; o_seen = 1; break; case 'n': is_by_qname = 1; break; case 'm': { char *q; max_mem = strtol(optarg, &q, 0); if (*q == 'k' || *q == 'K') max_mem <<= 10; else if (*q == 'm' || *q == 'M') max_mem <<= 20; else if (*q == 'g' || *q == 'G') max_mem <<= 30; break; } case 'T': kputs(optarg, &tmpprefix); break; case '@': n_threads = atoi(optarg); break; case 'l': level = atoi(optarg); break; default: if (parse_sam_global_opt(c, optarg, lopts, &ga) == 0) break; /* else fall-through */ case '?': sort_usage(stderr); ret = EXIT_FAILURE; goto sort_end; } } nargs = argc - optind; if (nargs == 0 && isatty(STDIN_FILENO)) { sort_usage(stdout); ret = EXIT_SUCCESS; goto sort_end; } else if (nargs >= 2) { // If exactly two, user probably tried to specify legacy if (nargs == 2) fprintf(stderr, "[bam_sort] Use -T PREFIX / -o FILE to specify temporary and final output files\n"); sort_usage(stderr); ret = EXIT_FAILURE; goto sort_end; } strcpy(modeout, "wb"); sam_open_mode(modeout+1, fnout, NULL); if (level >= 0) sprintf(strchr(modeout, '\0'), "%d", level < 9? level : 9); if (tmpprefix.l == 0) ksprintf(&tmpprefix, "%s.tmp", (nargs > 0)? argv[optind] : "STDIN"); ret = bam_sort_core_ext(is_by_qname, (nargs > 0)? argv[optind] : "-", tmpprefix.s, fnout, modeout, max_mem, n_threads, &ga.in, &ga.out); if (ret >= 0) ret = EXIT_SUCCESS; else { char dummy[4]; // If we failed on opening the input file & it has no .bam/.cram/etc // extension, the user probably tried legacy -o if (ret == -2 && o_seen && nargs > 0 && sam_open_mode(dummy, argv[optind], NULL) < 0) fprintf(stderr, "[bam_sort] Note the argument has been replaced by -T/-o options\n"); ret = EXIT_FAILURE; } sort_end: free(tmpprefix.s); sam_global_args_free(&ga); return ret; }