Denovogear trio - generates empty vcf file

[stanedav]

Hello, I I work on trios and I am trying to use denovogear. In our lab we are using two softwares for analysis - Surecall and NextGENe. I am able to make denovogear output from bam files generated in Surecall. But I have problem with NextGENe files, after running denovogear I will get empty vcf file and also in command line no progress is seen. My workflow is: 1) BAM files generated by NextGene (bam header is in file header1.txt) 2) samtools is used for reheader bam files - changed ID and SM in RG line to sample number (because all nextgene bam files has same ID and SM in header) code for reheading bam files: samtools reheader header.sam file.bam > newfile.bam (new header is in file header2.txt) 3) three bam files are merged into one bcf file with command: samtools mpileup -gDf hg19.fa father.bam mother.bam child.bam > merged.bcf 4) Denovogear analysis is performed by this command: ./dng dnm auto --ped fam6752.ped --bcf merged.bcf --output_vcf out.vcf (ped file is in 6752_fam.txt, I am using ID's from edited bam headers) Then, denovogear performs some analysis, but there is no progress in command line, and after few minutes it show, that analysis was performed with 0 results, here is output from analysis: DeNovoGear v1.1.1 posterior probability cutoff: 0 read depth filter: 0 output vcf file: /home/lab/galaxy-Win-Lin/David/trio/NG_data/6759_fam/6759.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 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 !

I tested two versions of Denovogear - 0.5.4 and 1.1.1, both with same results Also analysis with parameters: --pp_cutoff 0 and rd_cutoff 0 was performed, but with same results. Maybe there could be problem with wrong pairing between ped file and bam files, or am I doing something wrong?

Thank you for your advices

D. Stanek

Files: header1.txt header2.txt 6752_fam.txt

[reedacartwright]

@kaeldai what do you think?

[kaeldai]

It looks like DNM is not finding any snps or indels to analyze. This can happen if there's not any valid reference or alternative alleles at any sites.

Would it be possible to see some of the merged data in vcf format? The header and the first few sites should be good enough.