Having trouble getting non-empty output
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I'm trying to run denovogear on a WGS trio, and I'm not getting any variants in the output file. I'd appreciate any suggestions of what I may be doing wrong.
Our samples are called WG0840 (proband), WG0841 (mother), and WG0842 (father), and they were sequenced with 100bp paired-end reads with mean read depth of 50X on a BGISeq-500. The reads were aligned with BWA MEM (version 0.7.15), and processed with Picard (version 2.7.1, FixMateInformation, MarkDuplicates), and GATK (version 3.7, RealignerTargetCreator, IndelRealigner). I have denovogear version 1.1.1, and samtools 1.9. I'm running the command:
samtools mpileup -r 1:170120000-170140000 -f human_g1k_v37.fasta -gD WG084[012].bam | denovogear-v1.1.1-Linux-x86_64/bin/dng dnm auto --bcf - --pp_cutoff 0.0001 --ped WG0840-WG0841-WG0842_2.ped --output_vcf WG0840-WG0841-WG0842_denovogear_samtools.vcf
and WG0840-WG0841-WG0842_2.ped contains just one line:
WG0840 WG0840 WG0842 WG0841 1 1
I receive the following output on the console:
DeNovoGear v1.1.1
posterior probability cutoff: 0.0001
output vcf file: WG0840-WG0841-WG0842_denovogear_samtools.vcf
Created SNP lookup table
First mrate: 1 last: 1
First code: 6 last: 6
First target string: AA/AA/AA last: TT/TT/TT
First tref: 0.0002388 last: 0.99301
Created indel lookup table First code: 6 last: 6
First target string: RR/RR/RR last: DD/DD/DD
First prior: 0.05 last: 0.114
Created paired lookup table
First target string: AA/AA last: TT/TT
First prior 1 last: 1
[warning] samtools mpileup option `g` is functional, but deprecated. Please switch to using bcftools mpileup in future.
[warning] samtools mpileup option `D` is functional, but deprecated. Please switch to using bcftools mpileup in future.
[mpileup] 3 samples in 3 input files
Total number of SNP sites interrogated: 0
Total number of SNP sites passing read-depth filters: 0
Total number of INDEL sites interrogated: 0
Total number of INDEL sites passing read-depth filters: 0
Total number of Paired sample sites interrogated: 0
Total number of Paired sample sites passing read-depth filters: 0
Done !
But the vcf file does not contain any variants.
I know I have a de novo variant at 1:170,129,733, and it shows up very nicely in the samtools mpileup, but doesn't appear in the output file from denovogear. If I run:
samtools mpileup -r 1:170129733-170129735 -f human_g1k_v37.fasta r03_assembly/WG084[012].bam
then I get the following output:
1 170129733 C 43 ...,..,,,.,,,,,,.,..,,,,,..,.,,,,..,,,...,^]. BFiGFGFFEFFF@FFEFFGDFDGEGFGFFFFFFFGF?G=FFFE 47 ...,.,,,,......,.,.,,,....,,......,..,,.,,..,,. lFFFFFFFEFFFGFGGlGEFFFFFGFGFF=GFFGFFFGFCBFDCFFE 55 ,$,......,.,.,......,.......,....,...,,,...,,.,,,..,,.,^]. >FFFGGFFFFFFF;GGEmDAGFGFlFGBFF?G?EGGFGFGGFGFEEBF@G;FGEE
1 170129734 A 42 .GgGGgg,.,,gg,g.,G.gg,,,.G,G,,ggGG,g,G..g. FkGFFFFEFFFBGFFFF?FGDGFEFGFEFFGBFFEFFDFGFF 47 .$.$.,.,,,,......,.,.,,,....,,......,..,,.,,..,,. c;FFFFFFE1F>:FFFlFECFFGFGFGEFDFFGAFFFGF>AFEE@FE 56 ,......,.,.,.T....,.......,....,.,..,,,...,,.,,,..,,.,.^]. FFEDFFFFE7FFFECGlE9EFDFkFEFGGGFFE@GFFFFFFFGGGFF@FFGFFEGB
1 170129735 A 44 ..,..,,,.,,,,,,.,..,,,,,..,.,,,,..,,,...,..^]. FkEFFFFFFFF?GFFFFFFFFGFFFFFD=FCGFFFFFCFFGF7E 46 .,.,,,,......,.,.,,,....,,.....,.,..,,.,,..,,. FFCFFFFFBF?EFGkGFGFFFGFFEEFFFFFEGGFFGFC@FF8FFF 56 ,......,.,.,.T....,.......,....,.,..,,,...,,.,,,..,,.,.. GFDGFDF:FAFFEDFFk?EFFGFhAFGFFFGGF4FFFEFFFFFGFFFFFFGFFFFE
Any suggestions would be appreciated.
dng dnm is no longer supported because it was developed for an older version of samtools.