#include #include #include #ifdef HAVE_CONFIG_H #include "config.h" #endif #ifdef HAVE_PTHREAD #include #endif #include "bntseq.h" #include "bwt_lite.h" #include "utils.h" #include "bwtsw2.h" #include "stdaln.h" #include "kstring.h" #include "kseq.h" KSEQ_INIT(gzFile, gzread) #include "ksort.h" #define __left_lt(a, b) ((a).end > (b).end) KSORT_INIT(hit, bsw2hit_t, __left_lt) extern unsigned char nst_nt4_table[256]; unsigned char nt_comp_table[256] = { 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','T','V','G', 'H','N','N','C', 'D','N','N','M', 'N','K','N','N', 'N','N','Y','S', 'A','N','B','W', 'X','R','N','N', 'N','N','N','N', 'n','t','v','g', 'h','n','n','c', 'd','n','n','m', 'n','k','n','n', 'n','n','y','s', 'a','n','b','w', 'x','r','n','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N', 'N','N','N','N' }; extern int bsw2_resolve_duphits(const bwt_t *bwt, bwtsw2_t *b, int IS); extern int bsw2_resolve_query_overlaps(bwtsw2_t *b, float mask_level); bsw2opt_t *bsw2_init_opt() { bsw2opt_t *o = (bsw2opt_t*)calloc(1, sizeof(bsw2opt_t)); o->a = 1; o->b = 3; o->q = 5; o->r = 2; o->t = 30; o->bw = 50; o->z = 1; o->is = 3; o->t_seeds = 5; o->hard_clip = 0; o->mask_level = 0.50f; o->yita = 5.5f; o->coef = 5.5f; o->qr = o->q + o->r; o->n_threads = 1; o->chunk_size = 10000000; return o; } void bsw2_destroy(bwtsw2_t *b) { int i; if (b == 0) return; if (b->cigar) for (i = 0; i < b->n; ++i) free(b->cigar[i]); free(b->cigar); free(b->n_cigar); free(b->hits); free(b); } #define __gen_ap(par, opt) do { \ int i; \ for (i = 0; i < 25; ++i) (par).matrix[i] = -(opt)->b; \ for (i = 0; i < 4; ++i) (par).matrix[i*5+i] = (opt)->a; \ (par).gap_open = (opt)->q; (par).gap_ext = (opt)->r; \ (par).gap_end = (opt)->r; \ (par).row = 5; (par).band_width = opt->bw; \ } while (0) #define __rpac(pac, l, i) (pac[(l-i-1)>>2] >> (~(l-i-1)&3)*2 & 0x3) void bsw2_extend_left(const bsw2opt_t *opt, bwtsw2_t *b, uint8_t *_query, int lq, uint8_t *pac, uint32_t l_pac, int is_rev, uint8_t *_mem) { int i, matrix[25]; bwtint_t k; uint8_t *target = 0, *query; AlnParam par; par.matrix = matrix; __gen_ap(par, opt); query = calloc(lq, 1); // sort according to the descending order of query end ks_introsort(hit, b->n, b->hits); target = calloc(((lq + 1) / 2 * opt->a + opt->r) / opt->r + lq, 1); // reverse _query for (i = 0; i < lq; ++i) query[lq - i - 1] = _query[i]; // core loop for (i = 0; i < b->n; ++i) { bsw2hit_t *p = b->hits + i; int lt = ((p->beg + 1) / 2 * opt->a + opt->r) / opt->r + lq; int score, j; path_t path; p->n_seeds = 1; if (p->l || p->k == 0) continue; for (j = score = 0; j < i; ++j) { bsw2hit_t *q = b->hits + j; if (q->beg <= p->beg && q->k <= p->k && q->k + q->len >= p->k + p->len) { if (q->n_seeds < (1<<14) - 2) ++q->n_seeds; ++score; } } if (score) continue; if (lt > p->k) lt = p->k; if (is_rev) { for (k = p->k - 1, j = 0; k > 0 && j < lt; --k) // FIXME: k=0 not considered! target[j++] = __rpac(pac, l_pac, k); } else { for (k = p->k - 1, j = 0; k > 0 && j < lt; --k) // FIXME: k=0 not considered! target[j++] = pac[k>>2] >> (~k&3)*2 & 0x3; } lt = j; score = aln_extend_core(target, lt, query + lq - p->beg, p->beg, &par, &path, 0, p->G, _mem); if (score > p->G) { // extensible p->G = score; p->len += path.i; p->beg -= path.j; p->k -= path.i; } } free(query); free(target); } void bsw2_extend_rght(const bsw2opt_t *opt, bwtsw2_t *b, uint8_t *query, int lq, uint8_t *pac, uint32_t l_pac, int is_rev, uint8_t *_mem) { int i, matrix[25]; uint32_t k; uint8_t *target; AlnParam par; par.matrix = matrix; __gen_ap(par, opt); target = calloc(((lq + 1) / 2 * opt->a + opt->r) / opt->r + lq, 1); for (i = 0; i < b->n; ++i) { bsw2hit_t *p = b->hits + i; int lt = ((lq - p->beg + 1) / 2 * opt->a + opt->r) / opt->r + lq; int j, score; path_t path; if (p->l) continue; if (is_rev) { for (k = p->k, j = 0; k < p->k + lt && k < l_pac; ++k) target[j++] = __rpac(pac, l_pac, k); } else { for (k = p->k, j = 0; k < p->k + lt && k < l_pac; ++k) target[j++] = pac[k>>2] >> (~k&3)*2 & 0x3; } lt = j; score = aln_extend_core(target, lt, query + p->beg, lq - p->beg, &par, &path, 0, 1, _mem); // if (score < p->G) fprintf(stderr, "[bsw2_extend_hits] %d < %d\n", score, p->G); if (score >= p->G) { p->G = score; p->len = path.i; p->end = path.j + p->beg; } } free(target); } /* generate CIGAR array(s) in b->cigar[] */ static void gen_cigar(const bsw2opt_t *opt, int lq, uint8_t *seq[2], uint8_t *pac, bwtsw2_t *b) { uint8_t *target; int i, matrix[25]; AlnParam par; path_t *path; par.matrix = matrix; __gen_ap(par, opt); i = ((lq + 1) / 2 * opt->a + opt->r) / opt->r + lq; // maximum possible target length target = calloc(i, 1); path = calloc(i + lq, sizeof(path_t)); // memory clean up for b if (b->n < b->max) { b->max = b->n; b->hits = realloc(b->hits, b->n * sizeof(bsw2hit_t)); } if (b->cigar) free(b->cigar); if (b->n_cigar) free(b->n_cigar); b->cigar = (uint32_t**)calloc(b->max, sizeof(void*)); b->n_cigar = (int*)calloc(b->max, sizeof(int)); // generate CIGAR for (i = 0; i < b->n; ++i) { bsw2hit_t *p = b->hits + i; uint8_t *query; uint32_t k; int score, path_len, beg, end; if (p->l) continue; beg = (p->flag & 0x10)? lq - p->end : p->beg; end = (p->flag & 0x10)? lq - p->beg : p->end; query = seq[(p->flag & 0x10)? 1 : 0] + beg; for (k = p->k; k < p->k + p->len; ++k) // in principle, no out-of-boundary here target[k - p->k] = pac[k>>2] >> (~k&3)*2 & 0x3; score = aln_global_core(target, p->len, query, end - beg, &par, path, &path_len); b->cigar[i] = aln_path2cigar32(path, path_len, &b->n_cigar[i]); if (beg != 0 || end < lq) { // write soft clipping b->cigar[i] = realloc(b->cigar[i], 4 * (b->n_cigar[i] + 2)); if (beg != 0) { memmove(b->cigar[i] + 1, b->cigar[i], b->n_cigar[i] * 4); b->cigar[i][0] = beg<<4 | 4; ++b->n_cigar[i]; } if (end < lq) { b->cigar[i][b->n_cigar[i]] = (lq - end)<<4 | 4; ++b->n_cigar[i]; } } } free(target); free(path); } /* this is for the debugging purpose only */ void bsw2_debug_hits(const bwtsw2_t *b) { int i; printf("# raw hits: %d\n", b->n); for (i = 0; i < b->n; ++i) { bsw2hit_t *p = b->hits + i; if (p->l == 0) printf("%d, %d, %d, %u, %u\n", p->G, p->beg, p->end, p->k, p->l); } } static void merge_hits(bwtsw2_t *b[2], int l, int is_reverse) { int i; if (b[0]->n + b[1]->n > b[0]->max) { b[0]->max = b[0]->n + b[1]->n; b[0]->hits = realloc(b[0]->hits, b[0]->max * sizeof(bsw2hit_t)); } for (i = 0; i < b[1]->n; ++i) { bsw2hit_t *p = b[0]->hits + b[0]->n + i; *p = b[1]->hits[i]; if (is_reverse) { int x = p->beg; p->beg = l - p->end; p->end = l - x; p->flag |= 0x10; } } b[0]->n += b[1]->n; bsw2_destroy(b[1]); b[1] = 0; } /* seq[0] is the forward sequence and seq[1] is the reverse complement. */ static bwtsw2_t *bsw2_aln1_core(const bsw2opt_t *opt, const bntseq_t *bns, uint8_t *pac, const bwt_t *target, int l, uint8_t *seq[2], int is_rev, bsw2global_t *pool) { extern void bsw2_chain_filter(const bsw2opt_t *opt, int len, bwtsw2_t *b[2]); bwtsw2_t *b[2], **bb[2]; int k; for (k = 0; k < 2; ++k) { bwtl_t *query = bwtl_seq2bwtl(l, seq[k]); bb[k] = bsw2_core(opt, query, target, pool); bwtl_destroy(query); } b[0] = bb[0][1]; b[1] = bb[1][1]; // bb[*][1] are "narrow SA hits" bsw2_chain_filter(opt, l, b); for (k = 0; k < 2; ++k) { bsw2_extend_left(opt, bb[k][1], seq[k], l, pac, bns->l_pac, is_rev, pool->aln_mem); merge_hits(bb[k], l, 0); // bb[k][1] is merged to bb[k][0] here bsw2_resolve_duphits(0, bb[k][0], 0); bsw2_extend_rght(opt, bb[k][0], seq[k], l, pac, bns->l_pac, is_rev, pool->aln_mem); b[k] = bb[k][0]; free(bb[k]); } merge_hits(b, l, 1); // again, b[1] is merged to b[0] bsw2_resolve_query_overlaps(b[0], opt->mask_level); return b[0]; } /* set ->flag to records the origin of the hit (to forward bwt or reverse bwt) */ static void flag_fr(bwtsw2_t *b[2]) { int i, j; for (i = 0; i < b[0]->n; ++i) { bsw2hit_t *p = b[0]->hits + i; p->flag |= 0x10000; } for (i = 0; i < b[1]->n; ++i) { bsw2hit_t *p = b[1]->hits + i; p->flag |= 0x20000; } for (i = 0; i < b[0]->n; ++i) { bsw2hit_t *p = b[0]->hits + i; for (j = 0; j < b[1]->n; ++j) { bsw2hit_t *q = b[1]->hits + j; if (q->beg == p->beg && q->end == p->end && q->k == p->k && q->len == p->len && q->G == p->G) { q->flag |= 0x30000; p->flag |= 0x30000; break; } } } } typedef struct { int l, tid; char *name, *seq, *qual, *sam; } bsw2seq1_t; typedef struct { int n, max; bsw2seq1_t *seq; } bsw2seq_t; #ifdef HAVE_PTHREAD static pthread_mutex_t g_dbwtsw_lock = PTHREAD_MUTEX_INITIALIZER; #endif static int fix_cigar(const char *qname, const bntseq_t *bns, bsw2hit_t *p, int n_cigar, uint32_t *cigar) { // FIXME: this routine does not work if the query bridge three reference sequences int32_t coor, refl, lq; int x, y, i, seqid; bns_coor_pac2real(bns, p->k, p->len, &seqid); coor = p->k - bns->anns[seqid].offset; refl = bns->anns[seqid].len; x = coor; y = 0; // test if the alignment goes beyond the boundary for (i = 0; i < n_cigar; ++i) { int op = cigar[i]&0xf, ln = cigar[i]>>4; if (op == 1 || op == 4 || op == 5) y += ln; else if (op == 2) x += ln; else x += ln, y += ln; } lq = y; // length of the query sequence if (x > refl) { // then fix it int j, nc, mq[2], nlen[2]; uint32_t *cn, kk = 0; nc = mq[0] = mq[1] = nlen[0] = nlen[1] = 0; cn = calloc(n_cigar + 3, 4); x = coor; y = 0; for (i = j = 0; i < n_cigar; ++i) { int op = cigar[i]&0xf, ln = cigar[i]>>4; if (op == 4 || op == 5 || op == 1) { // ins or clipping y += ln; cn[j++] = cigar[i]; } else if (op == 2) { // del if (x + ln >= refl && nc == 0) { cn[j++] = (uint32_t)(lq - y)<<4 | 4; nc = j; cn[j++] = (uint32_t)y<<4 | 4; kk = p->k + (x + ln - refl); nlen[0] = x - coor; nlen[1] = p->len - nlen[0] - ln; } else cn[j++] = cigar[i]; x += ln; } else if (op == 0) { // match if (x + ln >= refl && nc == 0) { // FIXME: not consider a special case where a split right between M and I cn[j++] = (uint32_t)(refl - x)<<4 | 0; // write M cn[j++] = (uint32_t)(lq - y - (refl - x))<<4 | 4; // write S nc = j; mq[0] += refl - x; cn[j++] = (uint32_t)(y + (refl - x))<<4 | 4; if (x + ln - refl) cn[j++] = (uint32_t)(x + ln - refl)<<4 | 0; mq[1] += x + ln - refl; kk = bns->anns[seqid].offset + refl; nlen[0] = refl - coor; nlen[1] = p->len - nlen[0]; } else { cn[j++] = cigar[i]; mq[nc?1:0] += ln; } x += ln; y += ln; } } if (mq[0] > mq[1]) { // then take the first alignment n_cigar = nc; memcpy(cigar, cn, 4 * nc); p->len = nlen[0]; } else { p->k = kk; p->len = nlen[1]; n_cigar = j - nc; memcpy(cigar, cn + nc, 4 * (j - nc)); } free(cn); } return n_cigar; } /* generate SAM lines for a sequence in ks with alignment stored in * b. ks->name and ks->seq will be freed and set to NULL in the end. */ static void print_hits(const bntseq_t *bns, const bsw2opt_t *opt, bsw2seq1_t *ks, bwtsw2_t *b) { int i, k; kstring_t str; memset(&str, 0, sizeof(kstring_t)); if (b == 0 || b->n == 0) { // no hits ksprintf(&str, "%s\t4\t*\t0\t0\t*\t*\t0\t0\t", ks->name); for (i = 0; i < ks->l; ++i) kputc(ks->seq[i], &str); if (ks->qual) { kputc('\t', &str); for (i = 0; i < ks->l; ++i) kputc(ks->qual[i], &str); } else kputs("\t*", &str); kputc('\n', &str); } for (i = 0; b && i < b->n; ++i) { bsw2hit_t *p = b->hits + i; int32_t seqid = -1, coor = -1; int j, qual, nn = 0; int beg, end; if (p->l == 0) { b->n_cigar[i] = fix_cigar(ks->name, bns, p, b->n_cigar[i], b->cigar[i]); nn = bns_coor_pac2real(bns, p->k, p->len, &seqid); coor = p->k - bns->anns[seqid].offset; } ksprintf(&str, "%s\t%d", ks->name, p->flag&0x10); ksprintf(&str, "\t%s\t%d", seqid>=0? bns->anns[seqid].name : "*", coor + 1); if (p->l == 0) { { // estimate mapping quality float c = 1.0; int subo = p->G2 > opt->t? p->G2 : opt->t; if (p->flag>>16 == 1 || p->flag>>16 == 2) c *= .5; if (p->n_seeds < 2) c *= .2; qual = (int)(c * (p->G - subo) * (250.0 / p->G + 0.03 / opt->a) + .499); if (qual > 250) qual = 250; if (p->flag&1) qual = 0; } ksprintf(&str, "\t%d\t", qual); for (k = 0; k < b->n_cigar[i]; ++k) ksprintf(&str, "%d%c", b->cigar[i][k]>>4, (opt->hard_clip? "MIDNHHP" : "MIDNSHP")[b->cigar[i][k]&0xf]); } else ksprintf(&str, "\t0\t*"); ksprintf(&str, "\t*\t0\t0\t"); beg = 0; end = ks->l; if (opt->hard_clip) { if ((b->cigar[i][0]&0xf) == 4) beg += b->cigar[i][0]>>4; if ((b->cigar[i][b->n_cigar[i]-1]&0xf) == 4) end -= b->cigar[i][b->n_cigar[i]-1]>>4; } for (j = beg; j < end; ++j) { if (p->flag&0x10) kputc(nt_comp_table[(int)ks->seq[ks->l - 1 - j]], &str); else kputc(ks->seq[j], &str); } if (ks->qual) { kputc('\t', &str); for (j = beg; j < end; ++j) { if (p->flag&0x10) kputc(ks->qual[ks->l - 1 - j], &str); else kputc(ks->qual[j], &str); } } else ksprintf(&str, "\t*"); ksprintf(&str, "\tAS:i:%d\tXS:i:%d\tXF:i:%d\tXE:i:%d\tXN:i:%d", p->G, p->G2, p->flag>>16, p->n_seeds, nn); if (p->l) ksprintf(&str, "\tXI:i:%d", p->l - p->k + 1); kputc('\n', &str); } ks->sam = str.s; free(ks->seq); ks->seq = 0; free(ks->qual); ks->qual = 0; free(ks->name); ks->name = 0; } /* Core routine to align reads in _seq. It is separated from * process_seqs() to realize multi-threading */ static void bsw2_aln_core(int tid, bsw2seq_t *_seq, const bsw2opt_t *_opt, const bntseq_t *bns, uint8_t *pac, bwt_t * const target[2]) { int x; bsw2opt_t opt = *_opt; bsw2global_t *pool = bsw2_global_init(); for (x = 0; x < _seq->n; ++x) { bsw2seq1_t *p = _seq->seq + x; uint8_t *seq[2], *rseq[2]; int i, l, k; bwtsw2_t *b[2]; l = p->l; #ifdef HAVE_PTHREAD if (_opt->n_threads > 1) { pthread_mutex_lock(&g_dbwtsw_lock); if (p->tid < 0) p->tid = tid; else if (p->tid != tid) { pthread_mutex_unlock(&g_dbwtsw_lock); continue; } // in pinciple else should not happen pthread_mutex_unlock(&g_dbwtsw_lock); } #endif // set opt->t opt.t = _opt->t; if (opt.t < log(l) * opt.coef) opt.t = (int)(log(l) * opt.coef + .499); if (pool->max_l < l) { // then enlarge working space for aln_extend_core() int tmp = ((l + 1) / 2 * opt.a + opt.r) / opt.r + l; pool->max_l = l; pool->aln_mem = realloc(pool->aln_mem, (tmp + 2) * 24); } // set opt->bw opt.bw = _opt->bw; k = (l * opt.a - 2 * opt.q) / (2 * opt.r + opt.a); i = (l * opt.a - opt.a - opt.t) / opt.r; if (k > i) k = i; if (k < 1) k = 1; // I do not know if k==0 causes troubles opt.bw = _opt->bw < k? _opt->bw : k; // set seq[2] and rseq[2] seq[0] = calloc(l * 4, 1); seq[1] = seq[0] + l; rseq[0] = seq[1] + l; rseq[1] = rseq[0] + l; // convert sequences to 2-bit representation for (i = k = 0; i < l; ++i) { int c = nst_nt4_table[(int)p->seq[i]]; if (c >= 4) { c = (int)(drand48() * 4); ++k; } // FIXME: ambiguous bases are not properly handled seq[0][i] = c; seq[1][l-1-i] = 3 - c; rseq[0][l-1-i] = c; rseq[1][i] = 3 - c; } if (l - k < opt.t) { // too few unambiguous bases print_hits(bns, &opt, p, 0); free(seq[0]); continue; } // alignment b[0] = bsw2_aln1_core(&opt, bns, pac, target[0], l, seq, 0, pool); for (k = 0; k < b[0]->n; ++k) if (b[0]->hits[k].n_seeds < opt.t_seeds) break; if (k < b[0]->n) { b[1] = bsw2_aln1_core(&opt, bns, pac, target[1], l, rseq, 1, pool); for (i = 0; i < b[1]->n; ++i) { bsw2hit_t *p = b[1]->hits + i; int x = p->beg; p->beg = l - p->end; p->end = l - x; if (p->l == 0) p->k = bns->l_pac - (p->k + p->len); } flag_fr(b); merge_hits(b, l, 0); bsw2_resolve_duphits(0, b[0], 0); bsw2_resolve_query_overlaps(b[0], opt.mask_level); } else b[1] = 0; // generate CIGAR and print SAM gen_cigar(&opt, l, seq, pac, b[0]); print_hits(bns, &opt, p, b[0]); // free free(seq[0]); bsw2_destroy(b[0]); } bsw2_global_destroy(pool); } #ifdef HAVE_PTHREAD typedef struct { int tid; bsw2seq_t *_seq; const bsw2opt_t *_opt; const bntseq_t *bns; uint8_t *pac; bwt_t *target[2]; } thread_aux_t; /* another interface to bsw2_aln_core() to facilitate pthread_create() */ static void *worker(void *data) { thread_aux_t *p = (thread_aux_t*)data; bsw2_aln_core(p->tid, p->_seq, p->_opt, p->bns, p->pac, p->target); return 0; } #endif /* process sequences stored in _seq, generate SAM lines for these * sequences and reset _seq afterwards. */ static void process_seqs(bsw2seq_t *_seq, const bsw2opt_t *opt, const bntseq_t *bns, uint8_t *pac, bwt_t * const target[2]) { int i; #ifdef HAVE_PTHREAD if (opt->n_threads <= 1) { bsw2_aln_core(0, _seq, opt, bns, pac, target); } else { pthread_t *tid; pthread_attr_t attr; thread_aux_t *data; int j; pthread_attr_init(&attr); pthread_attr_setdetachstate(&attr, PTHREAD_CREATE_JOINABLE); data = (thread_aux_t*)calloc(opt->n_threads, sizeof(thread_aux_t)); tid = (pthread_t*)calloc(opt->n_threads, sizeof(pthread_t)); for (j = 0; j < opt->n_threads; ++j) { thread_aux_t *p = data + j; p->tid = j; p->_seq = _seq; p->_opt = opt; p->bns = bns; p->pac = pac; p->target[0] = target[0]; p->target[1] = target[1]; pthread_create(&tid[j], &attr, worker, p); } for (j = 0; j < opt->n_threads; ++j) pthread_join(tid[j], 0); free(data); free(tid); } #else bsw2_aln_core(0, _seq, opt, bns, pac, target); #endif // print and reset for (i = 0; i < _seq->n; ++i) { bsw2seq1_t *p = _seq->seq + i; if (p->sam) printf("%s", p->sam); free(p->name); free(p->seq); free(p->qual); free(p->sam); p->tid = -1; p->l = 0; p->name = p->seq = p->qual = p->sam = 0; } fflush(stdout); _seq->n = 0; } void bsw2_aln(const bsw2opt_t *opt, const bntseq_t *bns, bwt_t * const target[2], const char *fn) { gzFile fp; kseq_t *ks; int l, size = 0; uint8_t *pac; bsw2seq_t *_seq; pac = calloc(bns->l_pac/4+1, 1); if (pac == 0) { fprintf(stderr, "[bsw2_aln] insufficient memory!\n"); return; } for (l = 0; l < bns->n_seqs; ++l) printf("@SQ\tSN:%s\tLN:%d\n", bns->anns[l].name, bns->anns[l].len); fread(pac, 1, bns->l_pac/4+1, bns->fp_pac); fp = xzopen(fn, "r"); ks = kseq_init(fp); _seq = calloc(1, sizeof(bsw2seq_t)); while ((l = kseq_read(ks)) >= 0) { bsw2seq1_t *p; if (_seq->n == _seq->max) { _seq->max = _seq->max? _seq->max<<1 : 1024; _seq->seq = realloc(_seq->seq, _seq->max * sizeof(bsw2seq1_t)); } p = &_seq->seq[_seq->n++]; p->tid = -1; p->l = l; p->name = strdup(ks->name.s); p->seq = strdup(ks->seq.s); p->qual = ks->qual.l? strdup(ks->qual.s) : 0; p->sam = 0; size += l; if (size > opt->chunk_size) { fprintf(stderr, "[bsw2_aln] read %d sequences (%d bp)...\n", _seq->n, size); process_seqs(_seq, opt, bns, pac, target); size = 0; } } fprintf(stderr, "[bsw2_aln] read %d sequences (%d bp)...\n", _seq->n, size); process_seqs(_seq, opt, bns, pac, target); free(_seq->seq); free(_seq); kseq_destroy(ks); gzclose(fp); free(pac); }