#include #include #include "bam.h" #include "bam_maqcns.h" #include "ksort.h" #include "errmod.h" #include "kaln.h" KSORT_INIT_GENERIC(uint32_t) #define INDEL_WINDOW_SIZE 50 #define INDEL_EXT_DEP 0.9 typedef struct __bmc_aux_t { int max; uint32_t *info; uint16_t *info16; errmod_t *em; } bmc_aux_t; typedef struct { float esum[4], fsum[4]; uint32_t c[4]; } glf_call_aux_t; /* P() = \theta \sum_{i=1}^{N-1} 1/i P(D|) = \sum_{k=1}^{N-1} p_k 1/2 [(k/N)^n_2(1-k/N)^n_1 + (k/N)^n1(1-k/N)^n_2] p_k = 1/k / \sum_{i=1}^{N-1} 1/i */ static void cal_het(bam_maqcns_t *aa) { int k, n1, n2; double sum_harmo; // harmonic sum double poly_rate; free(aa->lhet); aa->lhet = (double*)calloc(256 * 256, sizeof(double)); sum_harmo = 0.0; for (k = 1; k <= aa->n_hap - 1; ++k) sum_harmo += 1.0 / k; for (n1 = 0; n1 < 256; ++n1) { for (n2 = 0; n2 < 256; ++n2) { long double sum = 0.0; double lC = aa->errmod == BAM_ERRMOD_SOAP? 0 : lgamma(n1+n2+1) - lgamma(n1+1) - lgamma(n2+1); for (k = 1; k <= aa->n_hap - 1; ++k) { double pk = 1.0 / k / sum_harmo; double log1 = log((double)k/aa->n_hap); double log2 = log(1.0 - (double)k/aa->n_hap); sum += pk * 0.5 * (expl(log1*n2) * expl(log2*n1) + expl(log1*n1) * expl(log2*n2)); } aa->lhet[n1<<8|n2] = lC + logl(sum); } } poly_rate = aa->het_rate * sum_harmo; aa->q_r = -4.343 * log(2.0 * poly_rate / (1.0 - poly_rate)); } /** initialize the helper structure */ static void cal_coef(bam_maqcns_t *aa) { int k, n, q; long double sum_a[257], b[256], q_c[256], tmp[256], fk2[256]; double *lC; if (aa->errmod == BAM_ERRMOD_MAQ2) return; // no need to do the following // aa->lhet will be allocated and initialized free(aa->fk); free(aa->coef); aa->coef = 0; aa->fk = (double*)calloc(256, sizeof(double)); aa->fk[0] = fk2[0] = 1.0; for (n = 1; n != 256; ++n) { aa->fk[n] = pow(aa->theta, n) * (1.0 - aa->eta) + aa->eta; fk2[n] = aa->fk[n>>1]; // this is an approximation, assuming reads equally likely come from both strands } if (aa->errmod == BAM_ERRMOD_SOAP) return; aa->coef = (double*)calloc(256*256*64, sizeof(double)); lC = (double*)calloc(256 * 256, sizeof(double)); for (n = 1; n != 256; ++n) for (k = 1; k <= n; ++k) lC[n<<8|k] = lgamma(n+1) - lgamma(k+1) - lgamma(n-k+1); for (q = 1; q != 64; ++q) { double e = pow(10.0, -q/10.0); double le = log(e); double le1 = log(1.0-e); for (n = 1; n != 256; ++n) { double *coef = aa->coef + (q<<16|n<<8); sum_a[n+1] = 0.0; for (k = n; k >= 0; --k) { // a_k = \sum_{i=k}^n C^n_k \epsilon^k (1-\epsilon)^{n-k} sum_a[k] = sum_a[k+1] + expl(lC[n<<8|k] + k*le + (n-k)*le1); b[k] = sum_a[k+1] / sum_a[k]; if (b[k] > 0.99) b[k] = 0.99; } for (k = 0; k != n; ++k) // log(\bar\beta_{nk}(\bar\epsilon)^{f_k}) q_c[k] = -4.343 * fk2[k] * logl(b[k] / e); for (k = 1; k != n; ++k) q_c[k] += q_c[k-1]; // \prod_{i=0}^k c_i for (k = 0; k <= n; ++k) { // powl() in 64-bit mode seems broken on my Mac OS X 10.4.9 tmp[k] = -4.343 * logl(1.0 - expl(fk2[k] * logl(b[k]))); coef[k] = (k? q_c[k-1] : 0) + tmp[k]; // this is the final c_{nk} } } } free(lC); } bam_maqcns_t *bam_maqcns_init() { bam_maqcns_t *bm; bm = (bam_maqcns_t*)calloc(1, sizeof(bam_maqcns_t)); bm->aux = (bmc_aux_t*)calloc(1, sizeof(bmc_aux_t)); bm->het_rate = 0.001; bm->theta = 0.83f; bm->n_hap = 2; bm->eta = 0.03; bm->cap_mapQ = 60; bm->min_baseQ = 13; return bm; } void bam_maqcns_prepare(bam_maqcns_t *bm) { if (bm->errmod == BAM_ERRMOD_MAQ2) bm->aux->em = errmod_init(1. - bm->theta); cal_coef(bm); cal_het(bm); } void bam_maqcns_destroy(bam_maqcns_t *bm) { if (bm == 0) return; free(bm->lhet); free(bm->fk); free(bm->coef); free(bm->aux->info); free(bm->aux->info16); if (bm->aux->em) errmod_destroy(bm->aux->em); free(bm->aux); free(bm); } glf1_t *bam_maqcns_glfgen(int _n, const bam_pileup1_t *pl, uint8_t ref_base, bam_maqcns_t *bm) { glf_call_aux_t *b = 0; int i, j, k, w[8], c, n; glf1_t *g = (glf1_t*)calloc(1, sizeof(glf1_t)); float p[16], min_p = 1e30; uint64_t rms; g->ref_base = ref_base; if (_n == 0) return g; // construct aux array if (bm->aux->max < _n) { bm->aux->max = _n; kroundup32(bm->aux->max); bm->aux->info = (uint32_t*)realloc(bm->aux->info, 4 * bm->aux->max); bm->aux->info16 = (uint16_t*)realloc(bm->aux->info16, 2 * bm->aux->max); } for (i = n = 0, rms = 0; i < _n; ++i) { const bam_pileup1_t *p = pl + i; uint32_t q, x = 0, qq; uint16_t y = 0; if (p->is_del || p->is_refskip || (p->b->core.flag&BAM_FUNMAP)) continue; q = (uint32_t)bam1_qual(p->b)[p->qpos]; if (q < bm->min_baseQ) continue; x |= (uint32_t)bam1_strand(p->b) << 18 | q << 8 | p->b->core.qual; y |= bam1_strand(p->b)<<4; if (p->b->core.qual < q) q = p->b->core.qual; c = p->b->core.qual < bm->cap_mapQ? p->b->core.qual : bm->cap_mapQ; rms += c * c; x |= q << 24; y |= q << 5; qq = bam1_seqi(bam1_seq(p->b), p->qpos); q = bam_nt16_nt4_table[qq? qq : ref_base]; if (!p->is_del && !p->is_refskip && q < 4) x |= 1 << 21 | q << 16, y |= q; bm->aux->info16[n] = y; bm->aux->info[n++] = x; } rms = (uint8_t)(sqrt((double)rms / n) + .499); if (bm->errmod == BAM_ERRMOD_MAQ2) { errmod_cal(bm->aux->em, n, 4, bm->aux->info16, p); goto goto_glf; } ks_introsort(uint32_t, n, bm->aux->info); // generate esum and fsum b = (glf_call_aux_t*)calloc(1, sizeof(glf_call_aux_t)); for (k = 0; k != 8; ++k) w[k] = 0; for (j = n - 1; j >= 0; --j) { // calculate esum and fsum uint32_t info = bm->aux->info[j]; if (info>>24 < 4 && (info>>8&0x3f) != 0) info = 4<<24 | (info&0xffffff); k = info>>16&7; if (info>>24 > 0) { b->esum[k&3] += bm->fk[w[k]] * (info>>24); b->fsum[k&3] += bm->fk[w[k]]; if (w[k] < 0xff) ++w[k]; ++b->c[k&3]; } } // rescale ->c[] for (j = c = 0; j != 4; ++j) c += b->c[j]; if (c > 255) { for (j = 0; j != 4; ++j) b->c[j] = (int)(254.0 * b->c[j] / c + 0.5); for (j = c = 0; j != 4; ++j) c += b->c[j]; } if (bm->errmod == BAM_ERRMOD_MAQ) { // generate likelihood for (j = 0; j != 4; ++j) { // homozygous float tmp1, tmp3; int tmp2, bar_e; for (k = 0, tmp1 = tmp3 = 0.0, tmp2 = 0; k != 4; ++k) { if (j == k) continue; tmp1 += b->esum[k]; tmp2 += b->c[k]; tmp3 += b->fsum[k]; } if (tmp2) { bar_e = (int)(tmp1 / tmp3 + 0.5); if (bar_e < 4) bar_e = 4; // should not happen if (bar_e > 63) bar_e = 63; p[j<<2|j] = tmp1 + bm->coef[bar_e<<16|c<<8|tmp2]; } else p[j<<2|j] = 0.0; // all the bases are j // heterozygous for (k = j + 1; k < 4; ++k) { for (i = 0, tmp2 = 0, tmp1 = tmp3 = 0.0; i != 4; ++i) { if (i == j || i == k) continue; tmp1 += b->esum[i]; tmp2 += b->c[i]; tmp3 += b->fsum[i]; } if (tmp2) { bar_e = (int)(tmp1 / tmp3 + 0.5); if (bar_e < 4) bar_e = 4; if (bar_e > 63) bar_e = 63; p[j<<2|k] = p[k<<2|j] = -4.343 * bm->lhet[b->c[j]<<8|b->c[k]] + tmp1 + bm->coef[bar_e<<16|c<<8|tmp2]; } else p[j<<2|k] = p[k<<2|j] = -4.343 * bm->lhet[b->c[j]<<8|b->c[k]]; // all the bases are either j or k } // for (k = 0; k != 4; ++k) if (p[j<<2|k] < 0.0) p[j<<2|k] = 0.0; } { // fix p[k<<2|k] float max1, max2, min1, min2; int max_k, min_k; max_k = min_k = -1; max1 = max2 = -1.0; min1 = min2 = 1e30; for (k = 0; k < 4; ++k) { if (b->esum[k] > max1) { max2 = max1; max1 = b->esum[k]; max_k = k; } else if (b->esum[k] > max2) max2 = b->esum[k]; } for (k = 0; k < 4; ++k) { if (p[k<<2|k] < min1) { min2 = min1; min1 = p[k<<2|k]; min_k = k; } else if (p[k<<2|k] < min2) min2 = p[k<<2|k]; } if (max1 > max2 && (min_k != max_k || min1 + 1.0 > min2)) p[max_k<<2|max_k] = min1 > 1.0? min1 - 1.0 : 0.0; } } else if (bm->errmod == BAM_ERRMOD_SOAP) { // apply the SOAP model // generate likelihood for (j = 0; j != 4; ++j) { float tmp; // homozygous for (k = 0, tmp = 0.0; k != 4; ++k) if (j != k) tmp += b->esum[k]; p[j<<2|j] = tmp; // heterozygous for (k = j + 1; k < 4; ++k) { for (i = 0, tmp = 0.0; i != 4; ++i) if (i != j && i != k) tmp += b->esum[i]; p[j<<2|k] = p[k<<2|j] = -4.343 * bm->lhet[b->c[j]<<8|b->c[k]] + tmp; } } } goto_glf: // convert necessary information to glf1_t g->ref_base = ref_base; g->max_mapQ = rms; g->depth = n > 16777215? 16777215 : n; for (j = 0; j != 4; ++j) for (k = j; k < 4; ++k) if (p[j<<2|k] < min_p) min_p = p[j<<2|k]; g->min_lk = min_p > 255.0? 255 : (int)(min_p + 0.5); for (j = c = 0; j != 4; ++j) for (k = j; k < 4; ++k) g->lk[c++] = p[j<<2|k]-min_p > 255.0? 255 : (int)(p[j<<2|k]-min_p + 0.5); free(b); return g; } uint32_t glf2cns(const glf1_t *g, int q_r) { int i, j, k, p[10], ref4; uint32_t x = 0; ref4 = bam_nt16_nt4_table[g->ref_base]; for (i = k = 0; i < 4; ++i) for (j = i; j < 4; ++j) { int prior = (i == ref4 && j == ref4? 0 : i == ref4 || j == ref4? q_r : q_r + 3); p[k] = (g->lk[k] + prior)<<4 | i<<2 | j; ++k; } for (i = 1; i < 10; ++i) // insertion sort for (j = i; j > 0 && p[j] < p[j-1]; --j) k = p[j], p[j] = p[j-1], p[j-1] = k; x = (1u<<(p[0]&3) | 1u<<(p[0]>>2&3)) << 28; // the best genotype x |= (uint32_t)g->max_mapQ << 16; // rms mapQ x |= ((p[1]>>4) - (p[0]>>4) < 256? (p[1]>>4) - (p[0]>>4) : 255) << 8; // consensus Q for (k = 0; k < 10; ++k) if ((p[k]&0xf) == (ref4<<2|ref4)) break; if (k == 10) k = 9; x |= (p[k]>>4) - (p[0]>>4) < 256? (p[k]>>4) - (p[0]>>4) : 255; // snp Q return x; } uint32_t bam_maqcns_call(int n, const bam_pileup1_t *pl, bam_maqcns_t *bm) { glf1_t *g; uint32_t x; if (n) { g = bam_maqcns_glfgen(n, pl, 0xf, bm); x = g->depth == 0? (0xfU<<28 | 0xfU<<24) : glf2cns(g, (int)(bm->q_r + 0.5)); free(g); } else x = 0xfU<<28 | 0xfU<<24; return x; } /************** *****************/ bam_maqindel_opt_t *bam_maqindel_opt_init() { bam_maqindel_opt_t *mi = (bam_maqindel_opt_t*)calloc(1, sizeof(bam_maqindel_opt_t)); mi->q_indel = 40; mi->r_indel = 0.00015; mi->r_snp = 0.001; // mi->mm_penalty = 3; mi->indel_err = 4; mi->ambi_thres = 10; return mi; } void bam_maqindel_ret_destroy(bam_maqindel_ret_t *mir) { if (mir == 0) return; free(mir->s[0]); free(mir->s[1]); free(mir); } int bam_tpos2qpos(const bam1_core_t *c, const uint32_t *cigar, int32_t tpos, int is_left, int32_t *_tpos) { int k, x = c->pos, y = 0, last_y = 0; *_tpos = c->pos; for (k = 0; k < c->n_cigar; ++k) { int op = cigar[k] & BAM_CIGAR_MASK; int l = cigar[k] >> BAM_CIGAR_SHIFT; if (op == BAM_CMATCH) { if (c->pos > tpos) return y; if (x + l > tpos) { *_tpos = tpos; return y + (tpos - x); } x += l; y += l; last_y = y; } else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) y += l; else if (op == BAM_CDEL || op == BAM_CREF_SKIP) { if (x + l > tpos) { *_tpos = is_left? x : x + l; return y; } x += l; } } *_tpos = x; return last_y; } #define MINUS_CONST 0x10000000 bam_maqindel_ret_t *bam_maqindel(int n, int pos, const bam_maqindel_opt_t *mi, const bam_pileup1_t *pl, const char *ref, int _n_types, int *_types) { int i, j, n_types, *types, left, right, max_rd_len = 0; bam_maqindel_ret_t *ret = 0; // if there is no proposed indel, check if there is an indel from the alignment if (_n_types == 0) { for (i = 0; i < n; ++i) { const bam_pileup1_t *p = pl + i; if (!(p->b->core.flag&BAM_FUNMAP) && p->indel != 0) break; } if (i == n) return 0; // no indel } { // calculate how many types of indels are available (set n_types and types) int m; uint32_t *aux; aux = (uint32_t*)calloc(n + _n_types + 1, 4); m = 0; aux[m++] = MINUS_CONST; // zero indel is always a type for (i = 0; i < n; ++i) { const bam_pileup1_t *p = pl + i; if (!(p->b->core.flag&BAM_FUNMAP) && p->indel != 0) aux[m++] = MINUS_CONST + p->indel; j = bam_cigar2qlen(&p->b->core, bam1_cigar(p->b)); if (j > max_rd_len) max_rd_len = j; } if (_n_types) // then also add this to aux[] for (i = 0; i < _n_types; ++i) if (_types[i]) aux[m++] = MINUS_CONST + _types[i]; ks_introsort(uint32_t, m, aux); // squeeze out identical types for (i = 1, n_types = 1; i < m; ++i) if (aux[i] != aux[i-1]) ++n_types; types = (int*)calloc(n_types, sizeof(int)); j = 0; types[j++] = aux[0] - MINUS_CONST; for (i = 1; i < m; ++i) { if (aux[i] != aux[i-1]) types[j++] = aux[i] - MINUS_CONST; } free(aux); } { // calculate left and right boundary left = pos > INDEL_WINDOW_SIZE? pos - INDEL_WINDOW_SIZE : 0; right = pos + INDEL_WINDOW_SIZE; if (types[0] < 0) right -= types[0]; // in case the alignments stand out the reference for (i = pos; i < right; ++i) if (ref[i] == 0) break; right = i; } { // the core part char *ref2, *rs, *inscns = 0; int qr_snp, k, l, *score, *pscore, max_ins = types[n_types-1]; qr_snp = (int)(-4.343 * log(mi->r_snp) + .499); if (max_ins > 0) { // get the consensus of inserted sequences int *inscns_aux = (int*)calloc(4 * n_types * max_ins, sizeof(int)); // count occurrences for (i = 0; i < n_types; ++i) { if (types[i] <= 0) continue; // not insertion for (j = 0; j < n; ++j) { const bam_pileup1_t *p = pl + j; if (!(p->b->core.flag&BAM_FUNMAP) && p->indel == types[i]) { for (k = 1; k <= p->indel; ++k) { int c = bam_nt16_nt4_table[bam1_seqi(bam1_seq(p->b), p->qpos + k)]; if (c < 4) ++inscns_aux[i*max_ins*4 + (k-1)*4 + c]; } } } } // construct the consensus of inserted sequence inscns = (char*)calloc(n_types * max_ins, sizeof(char)); for (i = 0; i < n_types; ++i) { for (j = 0; j < types[i]; ++j) { int max = 0, max_k = -1, *ia = inscns_aux + i*max_ins*4 + j*4; for (k = 0; k < 4; ++k) { if (ia[k] > max) { max = ia[k]; max_k = k; } } inscns[i*max_ins + j] = max? 1<= 0? -types[0] : -types[0] + types[i]; for (jj = 0; jj < tmp && j < right && ref[j]; ++jj, ++j) ref2[k++] = 4; } for (; j < right && ref[j]; ++j) ref2[k++] = bam_nt16_nt4_table[bam_nt16_table[(int)ref[j]]]; if (j < right) right = j; // calculate score for each read for (j = 0; j < n; ++j) { const bam_pileup1_t *p = pl + j; int qbeg, qend, tbeg, tend; if (p->b->core.flag & BAM_FUNMAP) continue; qbeg = bam_tpos2qpos(&p->b->core, bam1_cigar(p->b), left, 0, &tbeg); qend = bam_tpos2qpos(&p->b->core, bam1_cigar(p->b), right, 1, &tend); assert(tbeg >= left); for (l = qbeg; l < qend; ++l) rs[l - qbeg] = bam_nt16_nt4_table[bam1_seqi(bam1_seq(p->b), l)]; { int x, y, n_acigar, ps; uint32_t *acigar; ps = 0; if (tend - tbeg + types[i] <= 0) { score[i*n+j] = -(1<<20); pscore[i*n+j] = 1<<20; continue; } acigar = ka_global_core((uint8_t*)ref2 + tbeg - left, tend - tbeg + types[i], (uint8_t*)rs, qend - qbeg, &ap, &score[i*n+j], &n_acigar); x = tbeg - left; y = 0; for (l = 0; l < n_acigar; ++l) { int op = acigar[l]&0xf; int len = acigar[l]>>4; if (op == BAM_CMATCH) { int k; for (k = 0; k < len; ++k) if (ref2[x+k] != rs[y+k] && ref2[x+k] < 4) ps += bam1_qual(p->b)[y+k] < qr_snp? bam1_qual(p->b)[y+k] : qr_snp; x += len; y += len; } else if (op == BAM_CINS || op == BAM_CSOFT_CLIP) { if (op == BAM_CINS && l > 0 && l < n_acigar - 1) ps += mi->q_indel * len; y += len; } else if (op == BAM_CDEL) { if (l > 0 && l < n_acigar - 1) ps += mi->q_indel * len; x += len; } } pscore[i*n+j] = ps; /*if (1) { // for debugging only fprintf(stderr, "id=%d, pos=%d, type=%d, j=%d, score=%d, psore=%d, %d, %d, %d, %d, %d, ", j, pos+1, types[i], j, score[i*n+j], pscore[i*n+j], tbeg, tend, qbeg, qend, mi->q_indel); for (l = 0; l < n_acigar; ++l) fprintf(stderr, "%d%c", acigar[l]>>4, "MIDS"[acigar[l]&0xf]); fprintf(stderr, "\n"); for (l = 0; l < tend - tbeg + types[i]; ++l) fputc("ACGTN"[ref2[l+tbeg-left]], stderr); fputc('\n', stderr); for (l = 0; l < qend - qbeg; ++l) fputc("ACGTN"[rs[l]], stderr); fputc('\n', stderr); }*/ free(acigar); } } } { // get final result int *sum, max1, max2, max1_i, max2_i; // pick up the best two score sum = (int*)calloc(n_types, sizeof(int)); for (i = 0; i < n_types; ++i) for (j = 0; j < n; ++j) sum[i] += -pscore[i*n+j]; max1 = max2 = -0x7fffffff; max1_i = max2_i = -1; for (i = 0; i < n_types; ++i) { if (sum[i] > max1) { max2 = max1; max2_i = max1_i; max1 = sum[i]; max1_i = i; } else if (sum[i] > max2) { max2 = sum[i]; max2_i = i; } } free(sum); // write ret ret = (bam_maqindel_ret_t*)calloc(1, sizeof(bam_maqindel_ret_t)); ret->indel1 = types[max1_i]; ret->indel2 = types[max2_i]; ret->s[0] = (char*)calloc(abs(ret->indel1) + 2, 1); ret->s[1] = (char*)calloc(abs(ret->indel2) + 2, 1); // write indel sequence if (ret->indel1 > 0) { ret->s[0][0] = '+'; for (k = 0; k < ret->indel1; ++k) ret->s[0][k+1] = bam_nt16_rev_table[(int)inscns[max1_i*max_ins + k]]; } else if (ret->indel1 < 0) { ret->s[0][0] = '-'; for (k = 0; k < -ret->indel1 && ref[pos + k + 1]; ++k) ret->s[0][k+1] = ref[pos + k + 1]; } else ret->s[0][0] = '*'; if (ret->indel2 > 0) { ret->s[1][0] = '+'; for (k = 0; k < ret->indel2; ++k) ret->s[1][k+1] = bam_nt16_rev_table[(int)inscns[max2_i*max_ins + k]]; } else if (ret->indel2 < 0) { ret->s[1][0] = '-'; for (k = 0; k < -ret->indel2 && ref[pos + k + 1]; ++k) ret->s[1][k+1] = ref[pos + k + 1]; } else ret->s[1][0] = '*'; // write count for (i = 0; i < n; ++i) { const bam_pileup1_t *p = pl + i; if (p->indel == ret->indel1) ++ret->cnt1; else if (p->indel == ret->indel2) ++ret->cnt2; else ++ret->cnt_anti; } { // write gl[] int tmp, seq_err = 0; double x = 1.0; tmp = max1_i - max2_i; if (tmp < 0) tmp = -tmp; for (j = 0; j < tmp + 1; ++j) x *= INDEL_EXT_DEP; seq_err = mi->q_indel * (1.0 - x) / (1.0 - INDEL_EXT_DEP); ret->gl[0] = ret->gl[1] = 0; for (j = 0; j < n; ++j) { int s1 = pscore[max1_i*n + j], s2 = pscore[max2_i*n + j]; //fprintf(stderr, "id=%d, %d, %d, %d, %d, %d\n", j, pl[j].b->core.pos+1, types[max1_i], types[max2_i], s1, s2); if (s1 > s2) ret->gl[0] += s1 - s2 < seq_err? s1 - s2 : seq_err; else ret->gl[1] += s2 - s1 < seq_err? s2 - s1 : seq_err; } } // write cnt_ref and cnt_ambi if (max1_i != 0 && max2_i != 0) { for (j = 0; j < n; ++j) { int diff1 = score[j] - score[max1_i * n + j]; int diff2 = score[j] - score[max2_i * n + j]; if (diff1 > 0 && diff2 > 0) ++ret->cnt_ref; else if (diff1 == 0 || diff2 == 0) ++ret->cnt_ambi; } } } free(score); free(pscore); free(ref2); free(rs); free(inscns); } { // call genotype int q[3], qr_indel = (int)(-4.343 * log(mi->r_indel) + 0.5); int min1, min2, min1_i; q[0] = ret->gl[0] + (ret->s[0][0] != '*'? 0 : 0) * qr_indel; q[1] = ret->gl[1] + (ret->s[1][0] != '*'? 0 : 0) * qr_indel; q[2] = n * 3 + (ret->s[0][0] == '*' || ret->s[1][0] == '*'? 1 : 1) * qr_indel; min1 = min2 = 0x7fffffff; min1_i = -1; for (i = 0; i < 3; ++i) { if (q[i] < min1) { min2 = min1; min1 = q[i]; min1_i = i; } else if (q[i] < min2) min2 = q[i]; } ret->gt = min1_i; ret->q_cns = min2 - min1; // set q_ref if (ret->gt < 2) ret->q_ref = (ret->s[ret->gt][0] == '*')? 0 : q[1-ret->gt] - q[ret->gt] - qr_indel - 3; else ret->q_ref = (ret->s[0][0] == '*')? q[0] - q[2] : q[1] - q[2]; if (ret->q_ref < 0) ret->q_ref = 0; } free(types); return ret; }