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discover2.cpp
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/* The MIT License
Copyright (c) 2015 Adrian Tan <[email protected]>
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 "discover2.h"
namespace
{
/**
* For detecting overlapping reads.
*/
typedef struct
{
int32_t start1, end1;
} interval_t;
KHASH_MAP_INIT_STR(rdict, interval_t)
class Igor : Program
{
public:
///////////
//options//
///////////
std::vector<GenomeInterval> intervals;
std::string output_vcf_file;
std::string input_bam_file;
std::string ref_fasta_file;
std::string sample_id;
bool ignore_md;
int32_t debug;
//options for selecting reads
khash_t(rdict) *reads;
//sample properties
uint32_t ploidy;
//read filters
uint32_t read_mapq_cutoff;
uint16_t read_exclude_flag;
bool ignore_overlapping_read;
//variables for keeping track of chromosome
std::string chrom; //current chromosome
int32_t tid; // current sequence id in bam
int32_t rid; // current sequence id in bcf
///////
//i/o//
///////
BAMOrderedReader *odr;
bam1_t *s;
BCFOrderedWriter *odw;
bcf1_t *v;
/////////////////////////
//useful shared variables
/////////////////////////
int32_t *gt;
/////////
//stats//
/////////
uint32_t no_reads;
uint32_t no_overlapping_reads;
uint32_t no_passed_reads;
uint32_t no_exclude_flag_reads;
uint32_t no_low_mapq_reads;
uint32_t no_unaligned_cigars;
uint32_t no_malformed_del_cigars;
uint32_t no_malformed_ins_cigars;
uint32_t no_salvageable_ins_cigars;
uint32_t no_snps;
uint32_t no_ts;
uint32_t no_tv;
uint32_t no_insertions;
uint32_t no_deletions;
uint32_t no_left_soft_clips;
uint32_t no_right_soft_clips;
/////////
//tools//
/////////
Pileup pileup;
VariantFilter vf;
LogTool lt;
Igor(int argc, char **argv)
{
version = "0.5";
//////////////////////////
//options initialization//
//////////////////////////
try
{
std::string desc = "Discovers variants from reads in a SAM/BAM/CRAM file.";
TCLAP::CmdLine cmd(desc, ' ', version);
VTOutput my; cmd.setOutput(&my);
TCLAP::ValueArg<std::string> arg_intervals("i", "i", "intervals []", false, "", "str", cmd);
TCLAP::ValueArg<std::string> arg_interval_list("I", "I", "file containing list of intervals []", false, "", "file", cmd);
TCLAP::ValueArg<uint32_t> arg_debug("d", "d", "debug [0]", false, 0, "int", cmd);
TCLAP::SwitchArg arg_ignore_md("z", "z", "ignore MD tags [0]", cmd, false);
TCLAP::ValueArg<std::string> arg_output_vcf_file("o", "o", "output VCF file [-]", false, "-", "str", cmd);
TCLAP::ValueArg<std::string> arg_ref_fasta_file("r", "r", "reference sequence fasta file []", true, "", "str", cmd);
TCLAP::ValueArg<std::string> arg_sample_id("s", "s", "sample ID []", true, "", "str", cmd);
TCLAP::ValueArg<uint32_t> arg_ploidy("p", "p", "ploidy [2]", false, 2, "int", cmd);
//Reads
TCLAP::ValueArg<uint32_t> arg_read_mapq_cutoff("t", "t", "MAPQ cutoff for alignments (>) [20]", false, 20, "int", cmd);
TCLAP::SwitchArg arg_ignore_overlapping_read("l", "l", "ignore overlapping reads [false]", cmd, false);
TCLAP::ValueArg<uint32_t> arg_read_exclude_flag("a", "a", "read exclude flag [0x0704]", false, 0x0704, "int", cmd);
//Reference Bias
TCLAP::ValueArg<float> arg_reference_bias("B", "B", "reference bias [0.1]", false, 0.1, "float", cmd);
TCLAP::ValueArg<float> arg_lr_cutoff("C", "C", "likelihood ratio cutoff [0]", false, -1, "float", cmd);
//SNP
TCLAP::ValueArg<uint32_t> arg_snp_baseq_cutoff("q", "q", "base quality cutoff for bases [13]", false, 13, "int", cmd);
TCLAP::ValueArg<uint32_t> arg_snp_e_cutoff("e", "e", "snp evidence count cutoff [2]", false, 2, "int", cmd);
TCLAP::ValueArg<float> arg_snp_f_cutoff("f", "f", "snp fractional evidence cutoff [0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_snp_desired_type_I_error("j", "j", "snp desired type I error [0.0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_snp_desired_type_II_error("k", "k", "snp desired type II error [0.00001]", false, 0.00001, "float", cmd);
//DEL
TCLAP::ValueArg<uint32_t> arg_deletion_e_cutoff("u", "u", "deletion count cutoff [2]", false, 2, "int", cmd);
TCLAP::ValueArg<float> arg_deletion_f_cutoff("v", "v", "deletion fractional evidence cutoff [0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_deletion_desired_type_I_error("m", "m", "deletion desired type I error [0.0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_deletion_desired_type_II_error("n", "n", "deletion desired type II error [0.00001]", false, 0.00001, "float", cmd);
//INS
TCLAP::ValueArg<uint32_t> arg_insertion_e_cutoff("g", "g", "insertion count cutoff [2]", false, 2, "int", cmd);
TCLAP::ValueArg<float> arg_insertion_f_cutoff("h", "h", "insertion fractional evidence cutoff [0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_insertion_desired_type_I_error("c", "c", "insertion desired type I error [0.0]", false, 0, "float", cmd);
TCLAP::ValueArg<float> arg_insertion_desired_type_II_error("w", "w", "insertion desired type II error [0.00001]", false, 0.00001, "float", cmd);
//SOFT CLIPS
TCLAP::ValueArg<float> arg_sclip_mq_cutoff("x", "x", "soft clipped mean quality cutoff [0]", false, 0, "float", cmd);
TCLAP::ValueArg<uint32_t> arg_sclip_u_cutoff("y", "y", "soft clipped unique sequences cutoff [0]", false, 1, "float", cmd);
TCLAP::ValueArg<std::string> arg_input_bam_file("b", "b", "input SAM/BAM/CRAM file []", true, "", "string", cmd);
cmd.parse(argc, argv);
debug = arg_debug.getValue();
ignore_md = arg_ignore_md.getValue();
input_bam_file = arg_input_bam_file.getValue();
parse_intervals(intervals, arg_interval_list.getValue(), arg_intervals.getValue());
output_vcf_file = arg_output_vcf_file.getValue();
sample_id = arg_sample_id.getValue();
ploidy = arg_ploidy.getValue();
ref_fasta_file = arg_ref_fasta_file.getValue();
read_mapq_cutoff = arg_read_mapq_cutoff.getValue();
ignore_overlapping_read = arg_ignore_overlapping_read.getValue();
read_exclude_flag = arg_read_exclude_flag.getValue();
vf.set_reference_bias(arg_reference_bias.getValue());
vf.set_lr_cutoff(arg_lr_cutoff.getValue());
vf.set_snp_baseq_cutoff(arg_snp_baseq_cutoff.getValue());
vf.set_snp_e_cutoff(arg_snp_e_cutoff.getValue());
vf.set_snp_f_cutoff(arg_snp_f_cutoff.getValue());
vf.set_snp_desired_type_I_error(arg_snp_desired_type_I_error.getValue());
vf.set_snp_desired_type_II_error(arg_snp_desired_type_II_error.getValue());
vf.set_deletion_e_cutoff(arg_deletion_e_cutoff.getValue());
vf.set_deletion_f_cutoff(arg_deletion_f_cutoff.getValue());
vf.set_deletion_desired_type_I_error(arg_deletion_desired_type_I_error.getValue());
vf.set_deletion_desired_type_II_error(arg_deletion_desired_type_II_error.getValue());
vf.set_insertion_e_cutoff(arg_insertion_e_cutoff.getValue());
vf.set_insertion_f_cutoff(arg_insertion_f_cutoff.getValue());
vf.set_insertion_desired_type_I_error(arg_insertion_desired_type_I_error.getValue());
vf.set_insertion_desired_type_II_error(arg_insertion_desired_type_II_error.getValue());
vf.set_sclip_mq_cutoff(arg_sclip_mq_cutoff.getValue());
vf.set_sclip_u_cutoff(arg_sclip_u_cutoff.getValue());
vf.sync();
}
catch (TCLAP::ArgException &e)
{
std::cerr << "error: " << e.error() << " for arg " << e.argId() << "\n";
abort();
}
};
void initialize()
{
//////////////////////
//i/o initialization//
//////////////////////
//fails the following types
//1. unmapped reads
//2. secondary reads alignments
//3. failed QC filter
//4. duplicate
//read_exclude_flag = 0x0704;
odr = new BAMOrderedReader(input_bam_file, intervals, ref_fasta_file);
s = bam_init1();
odw = new BCFOrderedWriter(output_vcf_file, 0);
bam_hdr_transfer_contigs_to_bcf_hdr(odr->hdr, odw->hdr);
bcf_hdr_append(odw->hdr, "##ALT=<ID=RSC,Description=\"Right Soft Clip\">");
bcf_hdr_append(odw->hdr, "##ALT=<ID=LSC,Description=\"Left Soft Clip\">");
bcf_hdr_append(odw->hdr, "##INFO=<ID=SEQ,Number=1,Type=String,Description=\"Soft clipped Sequence\">");
bcf_hdr_append(odw->hdr, "##INFO=<ID=LLR,Number=1,Type=Float,Description=\"Log likelihood ratio: log10 [P(Non variant)/P(Variant)].\">");
bcf_hdr_append(odw->hdr, "##FORMAT=<ID=GT,Number=1,Type=String,Description=\"Genotype\">");
bcf_hdr_append(odw->hdr, "##FORMAT=<ID=E,Number=1,Type=Integer,Description=\"Number of reads containing evidence of the alternate allele\">");
bcf_hdr_append(odw->hdr, "##FORMAT=<ID=N,Number=1,Type=Integer,Description=\"Total number of reads at a candidate locus with reads that contain evidence of the alternate allele\">");
bcf_hdr_append(odw->hdr, "##FORMAT=<ID=MQS,Number=.,Type=Float,Description=\"Mean qualities of soft clipped bases.\">");
bcf_hdr_append(odw->hdr, "##FORMAT=<ID=STR,Number=.,Type=String,Description=\"Strands of soft clipped sequences.\">");
bcf_hdr_add_sample(odw->hdr, sample_id.c_str());
bcf_hdr_add_sample(odw->hdr, NULL);
v = bcf_init();
//for tracking overlapping reads
reads = kh_init(rdict);
////////////////////////
//stats initialization//
////////////////////////
no_reads = 0;
no_overlapping_reads = 0;
no_passed_reads = 0;
no_exclude_flag_reads = 0;
no_low_mapq_reads = 0;
no_unaligned_cigars = 0;
no_malformed_del_cigars = 0;
no_malformed_ins_cigars = 0;
no_salvageable_ins_cigars = 0;
no_snps = 0;
no_ts = 0;
no_tv = 0;
no_insertions = 0;
no_deletions = 0;
no_left_soft_clips = 0;
no_right_soft_clips = 0;
//////////////////////////////////////
//discovery variables initialization//
//////////////////////////////////////
chrom = "";
tid = -1;
rid = -1;
////////////////////////
//tools initialization//
////////////////////////
pileup.set_reference(ref_fasta_file);
pileup.set_debug(debug);
}
/**
* Print BAM for debugging purposes.
*/
void bam_print_key_values(bam_hdr_t *h, bam1_t *s)
{
const char* chrom = bam_get_chrom(h, s);
uint32_t pos1 = bam_get_pos1(s);
kstring_t seq = {0,0,0};
bam_get_seq_string(s, &seq);
uint32_t len = bam_get_l_qseq(s);
kstring_t qual = {0,0,0};
bam_get_qual_string(s, &qual);
kstring_t cigar_string = {0,0,0};
bam_get_cigar_string(s, &cigar_string);
kstring_t cigar_expanded_string = {0,0,0};
bam_get_cigar_expanded_string(s, &cigar_expanded_string);
uint16_t flag = bam_get_flag(s);
uint32_t mapq = bam_get_mapq(s);
uint8_t *aux;
char* md = NULL;
(aux=bam_aux_get(s, "MD")) && (md = bam_aux2Z(aux));
std::cerr << "##################" << "\n";
std::cerr << "chrom:pos: " << chrom << ":" << pos1 << "\n";
std::cerr << "read : " << seq.s << "\n";
std::cerr << "qual : " << qual.s << "\n";
std::cerr << "cigar_str: " << cigar_string.s << "\n";
std::cerr << "cigar : " << cigar_expanded_string.s << "\n";
std::cerr << "len : " << len << "\n";
std::cerr << "mapq : " << mapq << "\n";
std::cerr << "mpos1 : " << bam_get_mpos1(s) << "\n";
std::cerr << "mtid : " << bam_get_mtid(s) << "\n";
std::cerr << "md : " << (aux?md:"") << "\n";
std::cerr << "##################" << "\n";
if (seq.m) free(seq.s);
if (qual.m) free(qual.s);
if (cigar_string.m) free(cigar_string.s);
if (cigar_expanded_string.m) free(cigar_expanded_string.s);
}
/**
* Filter reads.
*
* Returns true if read is failed.
*/
bool filter_read(bam1_t *s)
{
khiter_t k;
int32_t ret;
if (ignore_overlapping_read)
{
//this read is part of a mate pair on the same contig
if (bam_get_mpos1(s) && (bam_get_tid(s)==bam_get_mtid(s)))
{
//first mate
if (bam_get_mpos1(s)>bam_get_pos1(s))
{
//overlapping
if (bam_get_mpos1(s)<=(bam_get_pos1(s) + bam_get_l_qseq(s) - 1))
{
//add read that has overlapping
//duplicate the record and perform the stitching later
char* qname = strdup(bam_get_qname(s));
k = kh_put(rdict, reads, qname, &ret);
if (!ret)
{
//already present
free(qname);
}
kh_val(reads, k) = {bam_get_pos1(s), bam_get_pos1(s)+bam_get_l_qseq(s)-1};
}
}
else
{
//check overlap
if((k = kh_get(rdict, reads, bam_get_qname(s)))!=kh_end(reads))
{
if (kh_exist(reads, k))
{
free((char*)kh_key(reads, k));
kh_del(rdict, reads, k);
++no_overlapping_reads;
}
//set this on to remove overlapping reads.
return false;
}
}
}
}
//should we join them up?
if(bam_get_flag(s) & read_exclude_flag)
{
//1. unmapped
//2. secondary alignment
//3. not passing QC
//4. PCR or optical duplicate
++no_exclude_flag_reads;
return false;
}
if (bam_get_mapq(s) < read_mapq_cutoff)
{
//filter short aligments and those with too many indels (?)
++no_low_mapq_reads;
return false;
}
//*****************************************************************
//should we have an assertion on the correctness of the bam record?
//Is, Ds not sandwiched in M
//leading and trailing Is - convert to S
//no Ms!!!!!
//*****************************************************************
int32_t n_cigar_op = bam_get_n_cigar_op(s);
if (n_cigar_op)
{
uint32_t *cigar = bam_get_cigar(s);
bool seenM = false;
int32_t last_opchr = '^';
for (int32_t i = 0; i < n_cigar_op; ++i)
{
int32_t opchr = bam_cigar_opchr(cigar[i]);
int32_t oplen = bam_cigar_oplen(cigar[i]);
if (opchr=='S')
{
if (i!=0 && i!=n_cigar_op-1)
{
std::cerr << "S issue\n";
bam_print_key_values(odr->hdr, s);
//++malformed_cigar;
}
}
else if (opchr=='M')
{
seenM = true;
}
else if (opchr=='D')
{
if (last_opchr!='M' || (i<=n_cigar_op && bam_cigar_opchr(cigar[i+1])!='M'))
{
std::cerr << "D issue\n";
++no_malformed_del_cigars;
bam_print_key_values(odr->hdr, s);
}
}
else if (opchr=='I')
{
if (last_opchr!='M' || (i<n_cigar_op && bam_cigar_opchr(cigar[i+1])!='M'))
{
if (last_opchr!='M')
{
if (last_opchr!='^' && last_opchr!='S')
{
std::cerr << "leading I issue\n";
bam_print_key_values(odr->hdr, s);
++no_malformed_ins_cigars;
}
else
{
++no_salvageable_ins_cigars;
}
}
else if (i==n_cigar_op-1)
{
++no_salvageable_ins_cigars;
}
else if (i==n_cigar_op-2 && (bam_cigar_opchr(cigar[i+1])=='S'))
{
++no_salvageable_ins_cigars;
}
else
{
std::cerr << "trailing I issue\n";
bam_print_key_values(odr->hdr, s);
++no_malformed_ins_cigars;
}
}
}
last_opchr = opchr;
}
if (!seenM)
{
std::cerr << "NO! M issue\n";
bam_print_key_values(odr->hdr, s);
++no_unaligned_cigars;
}
}
//check to see that hash should be cleared when encountering new contig.
//some bams may not be properly formed and contain orphaned sequences that
//can retained in the hash
if (bam_get_tid(s)!=tid)
{
for (k = kh_begin(reads); k != kh_end(reads); ++k)
{
if (kh_exist(reads, k))
{
free((char*)kh_key(reads, k));
kh_del(rdict, reads, k);
}
}
//tid is not updated here, it is handled by flush()
//kh_destroy(rdict, h);
}
return true;
}
/**
* Compute GLFSingle single variant likelihood ratio P(Non Variant)/P(Variant)
*/
double compute_glfsingle_llr(std::vector<uint32_t>& REF_Q, std::vector<uint32_t>& ALT_Q)
{
double pRR = 1;
double pRA = 1;
double pAA = 1;
double p;
double theta = 0.001;
for (uint32_t i=0; i<REF_Q.size(); ++i)
{
p = lt.pl2prob(REF_Q[i]);
pRR *= 1-p;
pRA *= 0.5;
pAA *= p;
}
for (uint32_t i=0; i<ALT_Q.size(); ++i)
{
p = lt.pl2prob(ALT_Q[i]);
pRR *= p;
pRA *= 0.5;
pAA *= 1-p;
}
return log10(pRR/((1-theta)*pRR+0.33*theta*pRA+0.67*theta*pAA));
}
/**
* Write out pileupPosition as a VCF entry if it contains a variant.
*/
void write_to_vcf(uint32_t rid, uint32_t gpos1, PileupPosition& p)
{
int32_t gts[2] = {0x0002,0x0004};
if (p.R=='N' || p.R=='X')
{
return;
}
if (p.X[1]+p.X[2]+p.X[4]+p.X[8]+p.X[15]>p.E+p.N ||
p.D.size()+p.I.size()>p.N)
{
std::cerr << "*******************\n";
std::cerr << "Evidence count ISSUES\n";
std::cerr << "#snp alts : " << (p.X[1]+p.X[2]+p.X[4]+p.X[8]+p.X[15]) << "\n";
std::cerr << "#del alts : " << p.D.size() << "\n";
std::cerr << "#ins alts : " << p.I.size() << "\n";
std::cerr << "#reads (N): " << p.N << "\n";
std::cerr << "#reads (E): " << p.E << "\n";
p.print(gpos1);
std::cerr << "*******************\n";
}
std::string alleles = "";
int32_t N = 0;
int32_t E = 0;
float LLR = 0;
kstring_t new_alleles = {0,0,0};
p.E = 0;
p.F = 0;
if (p.X[1])
{
if (vf.filter_snp(p.X[1], p.N+p.E-p.F))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, 'A');
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
E = p.X[1];
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
LLR = (float) compute_glfsingle_llr(p.REF_Q, p.ALT_Q);
bcf_update_info_float(odw->hdr, v, "LLR", &LLR, 1);
odw->write(v);
++no_snps;
if (p.R=='G')
{
++no_ts;
}
else
{
++no_tv;
}
}
}
if (p.X[2])
{
if (vf.filter_snp(p.X[2], p.N+p.E-p.F))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
v->unpacked = 1;
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, 'C');
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
E = p.X[2];
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
LLR = (float) compute_glfsingle_llr(p.REF_Q, p.ALT_Q);
bcf_update_info_float(odw->hdr, v, "LLR", &LLR, 1);
odw->write(v);
++no_snps;
if (p.R=='T')
{
++no_ts;
}
else
{
++no_tv;
}
}
}
if (p.X[4])
{
if (vf.filter_snp(p.X[4], p.N+p.E-p.F))
// if (vf.filter_snp(p.X[4], p.N+p.E))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, 'G');
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
E = p.X[4];
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
LLR = (float) compute_glfsingle_llr(p.REF_Q, p.ALT_Q);
bcf_update_info_float(odw->hdr, v, "LLR", &LLR, 1);
odw->write(v);
++no_snps;
if (p.R=='A')
{
++no_ts;
}
else
{
++no_tv;
}
}
}
if (p.X[8])
{
if (vf.filter_snp(p.X[8], p.N+p.E-p.F))
// if (vf.filter_snp(p.X[8], p.N+p.E))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, 'T');
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
E = p.X[8];
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
LLR = (float) compute_glfsingle_llr(p.REF_Q, p.ALT_Q);
bcf_update_info_float(odw->hdr, v, "LLR", &LLR, 1);
odw->write(v);
++no_snps;
if (p.R=='C')
{
++no_ts;
}
else
{
++no_tv;
}
}
}
if (false && p.X[15])
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, 'N');
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
E = p.X[15];
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
odw->write(v);
++no_snps;
}
if (p.D.size()!=0)
{
for (std::map<std::string, uint32_t>::iterator i = p.D.begin(); i!=p.D.end(); ++i)
{
const std::string& del = i->first;
if (del.find_first_of("Nn")==std::string::npos)
{
E = i->second;
N = p.N;
if (vf.filter_del(E, N))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(del);
alleles.append(1, ',');
alleles.append(1, p.R);
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
odw->write(v);
++no_deletions;
}
}
}
}
if (p.I.size()!=0)
{
for (std::map<std::string, uint32_t>::iterator i = p.I.begin(); i!=p.I.end(); ++i)
{
const std::string& ins = i->first;
if (ins.find_first_of("Nn")==std::string::npos)
{
E = i->second;
N = p.N;
if (vf.filter_ins(E, N))
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
alleles.clear();
alleles.append(1, p.R);
alleles.append(1, ',');
alleles.append(1, p.R);
alleles.append(ins);
bcf_update_alleles_str(odw->hdr, v, alleles.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
odw->write(v);
++no_insertions;
}
}
}
}
if (p.J.size()!=0)
{
if (false && p.J.size()>=vf.get_sclip_u_cutoff())
{
for (std::map<std::string, SoftClipInfo>::iterator i = p.J.begin(); i!=p.J.end(); ++i)
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
new_alleles.l = 0;
kputc(p.R, &new_alleles);
kputc(',', &new_alleles);
kputs("<LSC:", &new_alleles);
kputw(i->first.size(), &new_alleles);
kputc('>', &new_alleles);
bcf_update_alleles_str(odw->hdr, v, new_alleles.s);
bcf_update_info_string(odw->hdr, v, "SEQ", i->first.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
SoftClipInfo& info = i->second;
uint32_t no = info.no;
E = no;
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
bcf_update_format_float(odw->hdr, v, "MQS", &info.mean_quals[0], no);
bcf_update_format_char(odw->hdr, v, "STR", &info.strands[0], no);
odw->write(v);
++no_left_soft_clips;
}
}
}
if (p.K.size()!=0)
{
if (false && p.K.size()>=vf.get_sclip_u_cutoff())
{
for (std::map<std::string, SoftClipInfo>::iterator i = p.K.begin(); i!=p.K.end(); ++i)
{
bcf_clear(v);
bcf_set_rid(v, rid);
bcf_set_pos1(v, gpos1);
bcf_set_n_sample(v, 1);
new_alleles.l = 0;
kputc(p.R, &new_alleles);
kputc(',', &new_alleles);
kputs("<RSC:", &new_alleles);
kputw(i->first.size(), &new_alleles);
kputc('>', &new_alleles);
bcf_update_alleles_str(odw->hdr, v, new_alleles.s);
bcf_update_info_string(odw->hdr, v, "SEQ", i->first.c_str());
bcf_update_genotypes(odw->hdr, v, >s, ploidy);
SoftClipInfo& info = i->second;
uint32_t no = info.no;
E = no;
bcf_update_format_int32(odw->hdr, v, "E", &E, 1);
N = p.N+p.E;
bcf_update_format_int32(odw->hdr, v, "N", &N, 1);
float* mean_quals = info.mean_quals.data();
bcf_update_format_float(odw->hdr, v, "MQS", mean_quals, no);
char* s = info.strands.data();
bcf_update_format_char(odw->hdr, v, "STR", s, no);
odw->write(v);
++no_right_soft_clips;
}
}
}
if (new_alleles.m) free(new_alleles.s);
}
/**
* Check if flushable.
*
* returns
* 0 - not flushable
* 1 - flushable
* -1 - flushable, must update with new sequence
*/
int32_t flushable(bam1_t* s)
{
//different sequence, flush everything.
if (pileup.get_tid()!=bam_get_tid(s))
{
return -1;
}
//position is smaller than window size
if (pileup.get_window_size()>=bam_get_pos1(s))
{
return 0;
}
//some leeway for flushing the records
if (bam_get_pos1(s)-pileup.get_window_size()>pileup.get_gbeg1())
{
return 1;
}
else
{
return 0;
}
}
/**
* Flush records out before gpos1.
*/
void flush(bam1_t* s)
{
int32_t ret = 0;
if ((ret=flushable(s)))
{
if (ret==1)
{
if (debug>=3) std::cerr << "FLUSHING " << pileup.get_gbeg1() << " to " << bam_get_pos1(s)-pileup.get_window_size() << "\n";
uint32_t cpos1 = pileup.get_gbeg1();
uint32_t gpos1 = bam_get_pos1(s)-pileup.get_window_size();
uint32_t lend0 = pileup.get_gend1()<gpos1 ? pileup.end() : pileup.g2i(gpos1);
// uint32_t j;
// for (j=pileup.end(); j!=pileup.begin(); j=pileup.inc(j,1))
// {
// if(!pileup[j].is_cleared())
// {
// std::cerr << "NOT VALID!!!!\n";
// pileup[j].print();
// }
// }
uint32_t i;
for (i=pileup.begin(); i!=lend0; i=pileup.inc(i,1))
{
write_to_vcf(rid, cpos1, pileup[i]);
pileup[i].clear();
++cpos1;
}
pileup.set_gbeg1(cpos1);
pileup.set_beg0(i);
}
else //ret == -1
{
if (debug>=3) std::cerr << "FLUSHING " << pileup.get_gbeg1() << " to " << pileup.get_gend1() << "\n";
// uint32_t j;
// pileup.print_state();
// for (j=pileup.end(); j!=pileup.begin(); j=pileup.inc(j,1))
// {
// if(!pileup[j].is_cleared())
// {
// std::cerr << "NOT VALID!!!!\n";
// pileup[j].print();
// }
// }
uint32_t cpos1 = pileup.get_gbeg1();
uint32_t i;
for (i=pileup.begin(); i!=pileup.end(); i=pileup.inc(i,1))
{
write_to_vcf(rid, cpos1, pileup[i]);
pileup[i].clear();
++cpos1;
}
tid = bam_get_tid(s);
chrom.assign(bam_get_chrom(odr->hdr, s));
rid = bcf_hdr_name2id(odw->hdr, chrom.c_str());
pileup.set_tid(tid);
pileup.set_chrom(chrom);
pileup.set_gbeg1(0);
pileup.set_beg0(i);
}
}
}
/**
* Flush records till pileup is empty.
*/
void flush()
{
uint32_t cpos1 = pileup.get_gbeg1();
uint32_t i;
for (i=pileup.begin(); i!=pileup.end(); i=pileup.inc(i,1))
{
write_to_vcf(rid, cpos1, pileup[i]);
pileup[i].clear();
++cpos1;
}
tid = -1;
rid = -1;
pileup.set_tid(-1);
pileup.set_gbeg1(0);
pileup.set_beg0(0);
pileup.set_end0(0);