X-Git-Url: https://git.donarmstrong.com/?p=samtools.git;a=blobdiff_plain;f=bam2bcf.c;h=a51a406f249c05834b0d8e20f49403d5ed29a3b6;hp=1f7a0c042b2dad49abe5bb9d1668d90ddc96edf5;hb=307c147168f7154e3755712797078c513e0b242a;hpb=f27c5d351576e00cece3708adcb386da61fe7b7f diff --git a/bam2bcf.c b/bam2bcf.c index 1f7a0c0..a51a406 100644 --- a/bam2bcf.c +++ b/bam2bcf.c @@ -3,189 +3,229 @@ #include "bam.h" #include "kstring.h" #include "bam2bcf.h" +#include "errmod.h" #include "bcftools/bcf.h" -#define END_DIST_THRES 11 - extern void ks_introsort_uint32_t(size_t n, uint32_t a[]); #define CALL_ETA 0.03f #define CALL_MAX 256 -#define CALL_DEFTHETA 0.85f +#define CALL_DEFTHETA 0.83f +#define DEF_MAPQ 20 -struct __bcf_callaux_t { - int max_info, capQ, min_baseQ; - double *fk; - uint32_t *info; -}; +#define CAP_DIST 25 bcf_callaux_t *bcf_call_init(double theta, int min_baseQ) { bcf_callaux_t *bca; - int n; if (theta <= 0.) theta = CALL_DEFTHETA; bca = calloc(1, sizeof(bcf_callaux_t)); bca->capQ = 60; + bca->openQ = 40; bca->extQ = 20; bca->tandemQ = 100; bca->min_baseQ = min_baseQ; - bca->fk = calloc(CALL_MAX, sizeof(double)); - bca->fk[0] = 1.; - for (n = 1; n < CALL_MAX; ++n) - bca->fk[n] = theta >= 1.? 1. : pow(theta, n) * (1.0 - CALL_ETA) + CALL_ETA; - bca->fk[CALL_MAX-1] = 0.; + bca->e = errmod_init(1. - theta); + bca->min_frac = 0.002; + bca->min_support = 1; + bca->per_sample_flt = 0; return bca; } void bcf_call_destroy(bcf_callaux_t *bca) { - if (bca) { - free(bca->info); free(bca->fk); free(bca); - } + if (bca == 0) return; + errmod_destroy(bca->e); + free(bca->bases); free(bca->inscns); free(bca); } +/* ref_base is the 4-bit representation of the reference base. It is + * negative if we are looking at an indel. */ +int bcf_call_glfgen(int _n, const bam_pileup1_t *pl, int ref_base, bcf_callaux_t *bca, bcf_callret1_t *r) +{ + static int *var_pos = NULL, nvar_pos = 0; + int i, n, ref4, is_indel, ori_depth = 0; + memset(r, 0, sizeof(bcf_callret1_t)); + if (ref_base >= 0) { + ref4 = bam_nt16_nt4_table[ref_base]; + is_indel = 0; + } else ref4 = 4, is_indel = 1; + if (_n == 0) return -1; + // enlarge the bases array if necessary + if (bca->max_bases < _n) { + bca->max_bases = _n; + kroundup32(bca->max_bases); + bca->bases = (uint16_t*)realloc(bca->bases, 2 * bca->max_bases); + } + // fill the bases array + for (i = n = r->n_supp = 0; i < _n; ++i) { + const bam_pileup1_t *p = pl + i; + int q, b, mapQ, baseQ, is_diff, min_dist, seqQ; + // set base + if (p->is_del || p->is_refskip || (p->b->core.flag&BAM_FUNMAP)) continue; + ++ori_depth; + baseQ = q = is_indel? p->aux&0xff : (int)bam1_qual(p->b)[p->qpos]; // base/indel quality + seqQ = is_indel? (p->aux>>8&0xff) : 99; + if (q < bca->min_baseQ) continue; + if (q > seqQ) q = seqQ; + mapQ = p->b->core.qual < 255? p->b->core.qual : DEF_MAPQ; // special case for mapQ==255 + mapQ = mapQ < bca->capQ? mapQ : bca->capQ; + if (q > mapQ) q = mapQ; + if (q > 63) q = 63; + if (q < 4) q = 4; + if (!is_indel) { + b = bam1_seqi(bam1_seq(p->b), p->qpos); // base + b = bam_nt16_nt4_table[b? b : ref_base]; // b is the 2-bit base + is_diff = (ref4 < 4 && b == ref4)? 0 : 1; + } else { + b = p->aux>>16&0x3f; + is_diff = (b != 0); + } + if (is_diff) ++r->n_supp; + bca->bases[n++] = q<<5 | (int)bam1_strand(p->b)<<4 | b; + // collect annotations + if (b < 4) r->qsum[b] += q; + ++r->anno[0<<2|is_diff<<1|bam1_strand(p->b)]; + min_dist = p->b->core.l_qseq - 1 - p->qpos; + if (min_dist > p->qpos) min_dist = p->qpos; + if (min_dist > CAP_DIST) min_dist = CAP_DIST; + r->anno[1<<2|is_diff<<1|0] += baseQ; + r->anno[1<<2|is_diff<<1|1] += baseQ * baseQ; // FIXME: signed int is not enough for thousands of samples + r->anno[2<<2|is_diff<<1|0] += mapQ; + r->anno[2<<2|is_diff<<1|1] += mapQ * mapQ; // FIXME: signed int is not enough for thousands of samples + r->anno[3<<2|is_diff<<1|0] += min_dist; + r->anno[3<<2|is_diff<<1|1] += min_dist * min_dist; + } + r->depth = n; r->ori_depth = ori_depth; + // glfgen + errmod_cal(bca->e, n, 5, bca->bases, r->p); -typedef struct { - float esum[5], fsum[5]; - uint32_t c[5]; - int w[8]; -} auxaux_t; + // Calculate the Variant Distance Bias (make it optional?) + if ( nvar_pos < _n ) { + nvar_pos = _n; + var_pos = realloc(var_pos,sizeof(int)*nvar_pos); + } + int alt_dp=0, read_len=0; + for (i=0; i<_n; i++) { + const bam_pileup1_t *p = pl + i; + if ( bam1_seqi(bam1_seq(p->b),p->qpos) == ref_base ) + continue; -/* - The following code is nearly identical to bam_maqcns_glfgen() under - the simplified SOAPsnp model. It does the following: + var_pos[alt_dp] = p->qpos; + if ( (bam1_cigar(p->b)[0]&BAM_CIGAR_MASK)==4 ) + var_pos[alt_dp] -= bam1_cigar(p->b)[0]>>BAM_CIGAR_SHIFT; - 1) Collect strand, base, quality and mapping quality information for - each base and combine them in an integer: + alt_dp++; + read_len += p->b->core.l_qseq; + } + float mvd=0; + int j; + n=0; + for (i=0; imvd[0] = n ? mvd/n : 0; + r->mvd[1] = alt_dp; + r->mvd[2] = alt_dp ? read_len/alt_dp : 0; - x = min{baseQ,mapQ}<<24 | 1<<21 | strand<<18 | base<<16 | baseQ<<8 | mapQ + return r->depth; +} - 2) Sort the list of integers for the next step. - 3) For each base, calculate e_b, the sum of weighted qualities. For - each type of base on each strand, the best quality has the highest - weight. Only the top 255 bases on each strand are used (different - from maqcns). +void calc_vdb(int n, const bcf_callret1_t *calls, bcf_call_t *call) +{ + // Variant distance bias. Samples merged by means of DP-weighted average. - 4) Rescale the total read depth to 255. + float weight=0, tot_prob=0; - 5) Calculate Q(D|g) = -10\log_{10}P(D|g) (d_a is the allele count): + int i; + for (i=0; i)=\sum_{b\not=a}e_b - Q(D|)=3*(d_a+d_A)+\sum_{b\not=a,b\not=A}e_b - */ -int bcf_call_glfgen(int _n, const bam_pileup1_t *pl, int ref_base /*4-bit*/, bcf_callaux_t *bca, bcf_callret1_t *r) -{ - int i, j, k, c, n, ref4; - float *p = r->p; - auxaux_t aux; + if ( dp<2 ) continue; - memset(r, 0, sizeof(bcf_callret1_t)); - ref4 = bam_nt16_nt4_table[ref_base]; - if (_n == 0) return -1; + float prob = 0; + if ( dp==2 ) + { + // Exact formula + prob = (mvd==0) ? 1.0/read_len : (read_len-mvd)*2.0/read_len/read_len; + } + else if ( dp==3 ) + { + // Sin, quite accurate approximation + float mu = read_len/2.9; + prob = mvd>2*mu ? 0 : sin(mvd*3.14/2/mu) / (4*mu/3.14); + } + else + { + // Scaled gaussian curve, crude approximation, but behaves well. Using fixed depth for bigger depths. + if ( dp>5 ) + dp = 5; + float sigma2 = (read_len/1.9/(dp+1)) * (read_len/1.9/(dp+1)); + float norm = 1.125*sqrt(2*3.14*sigma2); + float mu = read_len/2.9; + if ( mvd < mu ) + prob = exp(-(mvd-mu)*(mvd-mu)/2/sigma2)/norm; + else + prob = exp(-(mvd-mu)*(mvd-mu)/3.125/sigma2)/norm; + } - // enlarge the aux array if necessary - if (bca->max_info < _n) { - bca->max_info = _n; - kroundup32(bca->max_info); - bca->info = (uint32_t*)realloc(bca->info, 4 * bca->max_info); - } - // fill the aux array - for (i = n = 0; i < _n; ++i) { - const bam_pileup1_t *p = pl + i; - uint32_t q, x = 0, qq; - int min_dist; - if (p->is_del || (p->b->core.flag&BAM_FUNMAP)) continue; // skip unmapped reads and deleted bases - q = (uint32_t)bam1_qual(p->b)[p->qpos]; // base quality - if (q < bca->min_baseQ) continue; - x |= (uint32_t)bam1_strand(p->b) << 18 | q << 8 | p->b->core.qual; - if (p->b->core.qual < q) q = p->b->core.qual; // cap the overall quality at mapping quality - x |= q << 24; - qq = bam1_seqi(bam1_seq(p->b), p->qpos); // base - q = bam_nt16_nt4_table[qq? qq : ref_base]; // q is the 2-bit base - if (q < 4) x |= 1 << 21 | q << 16; - k = (ref4 < 4 && q == ref4)? 0 : 1; - k = k<<1 | bam1_strand(p->b); - ++r->d[k]; - bca->info[n++] = x; - // calculate min_dist - min_dist = p->b->core.l_qseq - 1 - p->qpos; - if (min_dist > p->qpos) min_dist = p->qpos; - k = (k&2) | (min_dist <= END_DIST_THRES); - ++r->ed[k]; - } - ks_introsort_uint32_t(n, bca->info); - r->depth = n; - // generate esum and fsum - memset(&aux, 0, sizeof(auxaux_t)); - for (j = n - 1, r->sum_Q2 = 0; j >= 0; --j) { // calculate esum and fsum - uint32_t info = bca->info[j]; - int tmp; - if (info>>24 < 4 && (info>>8&0x3f) != 0) info = 4<<24 | (info&0xffffff); - k = info>>16&7; - if (info>>24 > 0) { - aux.esum[k&3] += bca->fk[aux.w[k]] * (info>>24); - aux.fsum[k&3] += bca->fk[aux.w[k]]; - if (aux.w[k] + 1 < CALL_MAX) ++aux.w[k]; - ++aux.c[k&3]; - } - tmp = (int)(info&0xff) < bca->capQ? (int)(info&0xff) : bca->capQ; - r->sum_Q2 += tmp * tmp; - } - memcpy(r->esum, aux.esum, 5 * sizeof(float)); - // rescale ->c[] - for (j = c = 0; j != 4; ++j) c += aux.c[j]; - if (c > 255) { - for (j = 0; j != 4; ++j) aux.c[j] = (int)(254.0 * aux.c[j] / c + 0.499); - for (j = c = 0; j != 4; ++j) c += aux.c[j]; - } - // generate likelihood - for (j = 0; j != 5; ++j) { - float tmp; - // homozygous - for (k = 0, tmp = 0.0; k != 5; ++k) - if (j != k) tmp += aux.esum[k]; - p[j*5+j] = tmp; // anything that is not j - // heterozygous - for (k = j + 1; k < 5; ++k) { - for (i = 0, tmp = 0.0; i != 5; ++i) - if (i != j && i != k) tmp += aux.esum[i]; - p[j*5+k] = p[k*5+j] = 3.01 * (aux.c[j] + aux.c[k]) + tmp; - } - } - return r->depth; + //fprintf(stderr,"dp=%d mvd=%d read_len=%d -> prob=%f\n", dp,mvd,read_len,prob); + tot_prob += prob*dp; + weight += dp; + } + tot_prob = weight ? tot_prob/weight : 1; + //fprintf(stderr,"prob=%f\n", tot_prob); + call->vdb = tot_prob; } int bcf_call_combine(int n, const bcf_callret1_t *calls, int ref_base /*4-bit*/, bcf_call_t *call) { - int ref4, i, j; - int64_t sum[5], tmp; - call->ori_ref = ref4 = bam_nt16_nt4_table[ref_base]; - if (ref4 > 4) ref4 = 4; - { // calculate esum - double esum[5]; - memset(esum, 0, sizeof(double) * 4); - for (i = 0; i < n; ++i) { - for (j = 0; j < 4; ++j) - esum[j] += calls[i].esum[j]; - } + int ref4, i, j, qsum[4]; + int64_t tmp; + if (ref_base >= 0) { + call->ori_ref = ref4 = bam_nt16_nt4_table[ref_base]; + if (ref4 > 4) ref4 = 4; + } else call->ori_ref = -1, ref4 = 0; + // calculate qsum + memset(qsum, 0, 4 * sizeof(int)); + for (i = 0; i < n; ++i) for (j = 0; j < 4; ++j) - sum[j] = (int)(esum[j] * 100. + .499) << 2 | j; - } + qsum[j] += calls[i].qsum[j]; + int qsum_tot=0; + for (j=0; j<4; j++) { qsum_tot += qsum[j]; call->qsum[j] = 0; } + for (j = 0; j < 4; ++j) qsum[j] = qsum[j] << 2 | j; // find the top 2 alleles for (i = 1; i < 4; ++i) // insertion sort - for (j = i; j > 0 && sum[j] < sum[j-1]; --j) - tmp = sum[j], sum[j] = sum[j-1], sum[j-1] = tmp; + for (j = i; j > 0 && qsum[j] < qsum[j-1]; --j) + tmp = qsum[j], qsum[j] = qsum[j-1], qsum[j-1] = tmp; // set the reference allele and alternative allele(s) for (i = 0; i < 5; ++i) call->a[i] = -1; call->unseen = -1; call->a[0] = ref4; for (i = 3, j = 1; i >= 0; --i) { - if ((sum[i]&3) != ref4) { - if (sum[i]>>2 != 0) call->a[j++] = sum[i]&3; + if ((qsum[i]&3) != ref4) { + if (qsum[i]>>2 != 0) + { + if ( j<4 ) call->qsum[j] = (float)(qsum[i]>>2)/qsum_tot; // ref N can make j>=4 + call->a[j++] = qsum[i]&3; + } else break; } + else + call->qsum[0] = (float)(qsum[i]>>2)/qsum_tot; + } + if (ref_base >= 0) { // for SNPs, find the "unseen" base + if (((ref4 < 4 && j < 4) || (ref4 == 4 && j < 5)) && i >= 0) + call->unseen = j, call->a[j++] = qsum[i]&3; + call->n_alleles = j; + } else { + call->n_alleles = j; + if (call->n_alleles == 1) return -1; // no reliable supporting read. stop doing anything } - if (((ref4 < 4 && j < 4) || (ref4 == 4 && j < 5)) && i >= 0) - call->unseen = j, call->a[j++] = sum[i]&3; - call->n_alleles = j; // set the PL array if (call->n < n) { call->n = n; @@ -197,8 +237,8 @@ int bcf_call_combine(int n, const bcf_callret1_t *calls, int ref_base /*4-bit*/, x = call->n_alleles * (call->n_alleles + 1) / 2; // get the possible genotypes for (i = z = 0; i < call->n_alleles; ++i) - for (j = i; j < call->n_alleles; ++j) - g[z++] = call->a[i] * 5 + call->a[j]; + for (j = 0; j <= i; ++j) + g[z++] = call->a[j] * 5 + call->a[i]; for (i = 0; i < n; ++i) { uint8_t *PL = call->PL + x * i; const bcf_callret1_t *r = calls + i; @@ -213,52 +253,110 @@ int bcf_call_combine(int n, const bcf_callret1_t *calls, int ref_base /*4-bit*/, PL[j] = y; } } +// if (ref_base < 0) fprintf(stderr, "%d,%d,%f,%d\n", call->n_alleles, x, sum_min, call->unseen); call->shift = (int)(sum_min + .499); } - memset(call->d, 0, 4 * sizeof(int)); - memset(call->ed, 0, 4 * sizeof(int)); - for (i = call->depth = 0, tmp = 0; i < n; ++i) { + // combine annotations + memset(call->anno, 0, 16 * sizeof(int)); + for (i = call->depth = call->ori_depth = 0, tmp = 0; i < n; ++i) { call->depth += calls[i].depth; - for (j = 0; j < 4; ++j) call->d[j] += calls[i].d[j], call->ed[j] += calls[i].ed[j]; - tmp += calls[i].sum_Q2; + call->ori_depth += calls[i].ori_depth; + for (j = 0; j < 16; ++j) call->anno[j] += calls[i].anno[j]; } - call->rmsQ = (int)(sqrt((double)tmp / call->depth) + .499); + + calc_vdb(n, calls, call); + return 0; } -int bcf_call2bcf(int tid, int pos, bcf_call_t *bc, bcf1_t *b) +int bcf_call2bcf(int tid, int pos, bcf_call_t *bc, bcf1_t *b, bcf_callret1_t *bcr, int fmt_flag, + const bcf_callaux_t *bca, const char *ref) { + extern double kt_fisher_exact(int n11, int n12, int n21, int n22, double *_left, double *_right, double *two); kstring_t s; - int i; + int i, j; + b->n_smpl = bc->n; b->tid = tid; b->pos = pos; b->qual = 0; s.s = b->str; s.m = b->m_str; s.l = 0; kputc('\0', &s); - kputc("ACGTN"[bc->ori_ref], &s); kputc('\0', &s); - for (i = 1; i < 5; ++i) { - if (bc->a[i] < 0) break; - if (i > 1) kputc(',', &s); -// kputc(bc->unseen == i && i != 3? 'X' : "ACGT"[bc->a[i]], &s); - kputc(bc->unseen == i? 'X' : "ACGT"[bc->a[i]], &s); + if (bc->ori_ref < 0) { // an indel + // write REF + kputc(ref[pos], &s); + for (j = 0; j < bca->indelreg; ++j) kputc(ref[pos+1+j], &s); + kputc('\0', &s); + // write ALT + kputc(ref[pos], &s); + for (i = 1; i < 4; ++i) { + if (bc->a[i] < 0) break; + if (i > 1) { + kputc(',', &s); kputc(ref[pos], &s); + } + if (bca->indel_types[bc->a[i]] < 0) { // deletion + for (j = -bca->indel_types[bc->a[i]]; j < bca->indelreg; ++j) + kputc(ref[pos+1+j], &s); + } else { // insertion; cannot be a reference unless a bug + char *inscns = &bca->inscns[bc->a[i] * bca->maxins]; + for (j = 0; j < bca->indel_types[bc->a[i]]; ++j) + kputc("ACGTN"[(int)inscns[j]], &s); + for (j = 0; j < bca->indelreg; ++j) kputc(ref[pos+1+j], &s); + } + } + kputc('\0', &s); + } else { // a SNP + kputc("ACGTN"[bc->ori_ref], &s); kputc('\0', &s); + for (i = 1; i < 5; ++i) { + if (bc->a[i] < 0) break; + if (i > 1) kputc(',', &s); + kputc(bc->unseen == i? 'X' : "ACGT"[bc->a[i]], &s); + } + kputc('\0', &s); } kputc('\0', &s); - kputc('\0', &s); // INFO - kputs("MQ=", &s); kputw(bc->rmsQ, &s); // kputs(";DP=", &s); kputw(bc->depth, &s); - kputs(";DP4=", &s); - for (i = 0; i < 4; ++i) { - if (i) kputc(',', &s); - kputw(bc->d[i], &s); - } - kputs(";ED4=", &s); - for (i = 0; i < 4; ++i) { + if (bc->ori_ref < 0) ksprintf(&s,"INDEL;IS=%d,%f;", bca->max_support, bca->max_frac); + kputs("DP=", &s); kputw(bc->ori_depth, &s); kputs(";I16=", &s); + for (i = 0; i < 16; ++i) { if (i) kputc(',', &s); - kputw(bc->ed[i], &s); + kputw(bc->anno[i], &s); } + ksprintf(&s,";QS=%f,%f,%f,%f", bc->qsum[0],bc->qsum[1],bc->qsum[2],bc->qsum[3]); + if (bc->vdb != 1) + ksprintf(&s, ";VDB=%.4f", bc->vdb); kputc('\0', &s); // FMT - kputs("PL", &s); kputc('\0', &s); + kputs("PL", &s); + if (bcr && fmt_flag) { + if (fmt_flag & B2B_FMT_DP) kputs(":DP", &s); + if (fmt_flag & B2B_FMT_DV) kputs(":DV", &s); + if (fmt_flag & B2B_FMT_SP) kputs(":SP", &s); + } + kputc('\0', &s); b->m_str = s.m; b->str = s.s; b->l_str = s.l; - bcf_sync(bc->n, b); + bcf_sync(b); memcpy(b->gi[0].data, bc->PL, b->gi[0].len * bc->n); + if (bcr && fmt_flag) { + uint16_t *dp = (fmt_flag & B2B_FMT_DP)? b->gi[1].data : 0; + uint16_t *dv = (fmt_flag & B2B_FMT_DV)? b->gi[1 + ((fmt_flag & B2B_FMT_DP) != 0)].data : 0; + int32_t *sp = (fmt_flag & B2B_FMT_SP)? b->gi[1 + ((fmt_flag & B2B_FMT_DP) != 0) + ((fmt_flag & B2B_FMT_DV) != 0)].data : 0; + for (i = 0; i < bc->n; ++i) { + bcf_callret1_t *p = bcr + i; + if (dp) dp[i] = p->depth < 0xffff? p->depth : 0xffff; + if (dv) dv[i] = p->n_supp < 0xffff? p->n_supp : 0xffff; + if (sp) { + if (p->anno[0] + p->anno[1] < 2 || p->anno[2] + p->anno[3] < 2 + || p->anno[0] + p->anno[2] < 2 || p->anno[1] + p->anno[3] < 2) + { + sp[i] = 0; + } else { + double left, right, two; + int x; + kt_fisher_exact(p->anno[0], p->anno[1], p->anno[2], p->anno[3], &left, &right, &two); + x = (int)(-4.343 * log(two) + .499); + if (x > 255) x = 255; + sp[i] = x; + } + } + } + } return 0; }