SNP calling in GATK

Question

[aj123]

java -jar /../../GenomeAnalysisTK.jar -T UnifiedGenotyper -R /data/../hg19_karyotypic.fa -I /../samplesorted.bam.bai -o samplesorted.geli.calls -vf GELI -stand_call_conf 30.0 -stand_emit_conf 10.0

I'm getting an error saying -vf argument not defined. and I should use -h argument. I have a sorted BAM file and just want to compare it to the reference genome and generate a vcf file. That's all. many thanks for your help.

[Pierre Lindenbaum]

I cannot see the option `-vf` in https://www.broadinstitute.org/gatk/gatkdocs/org_broadinstitute_gatk_tools_walkers_genotyper_UnifiedGenotyper.php

[aj123]

You are right, Pierre. Ive removed that I believe it was for an old version now the default output is only vcf files. Now im getting an error saying "strictness is silent".

[aj123]

You are right, Pierre. Ive removed that I believe it was for an old version now the default output is only vcf files.

I retried with

java -jar /data/odity/EXOME_AD_CCAMP-Batch2/02_0M_N/AD1/GenomeAnalysisTK-2.6-5-gba531bd/GenomeAnalysisTK.jar -R /data/odity/REF_GENOME_hg19/hg19_karyotypic.fa -T UnifiedGenotyper -I /data/odity/EXOME_AD_CCAMP-Batch2/02_0M_N/AD1/AD1sorted.bam -o outprefix.snps.vcf -stand_call_conf 50.0 -stand_emit_conf 10.0

Now I'm getting the error: The GATK no longer supports SAM files without read groups.

Answer 1

[Chirag Nepal]

Have a look at this tutorial

http://www.cureffi.org/2012/09/19/exome-sequencing-pipeline-using-gatk/

[aj123]

Thanks Chirag. Rectified the above error. Retried with

java -jar /../GenomeAnalysisTK-2.6-5-gba531bd/GenomeAnalysisTK.jar -R /data/../REF_GENOME_hg19/hg19_karyotypic.fa -T UnifiedGenotyper -I /data/../AD1cordsortrealigned.bam -o outprefix.snps.vcf -stand_call_conf 50.0 -stand_emit_conf 10.0

and now I'm getting:

##### ERROR MESSAGE: Input files reads and reference have incompatible contigs: Relative ordering of overlapping contigs differs, which is unsafe.
##### ERROR   reads contigs = [chr1, chr2, chr3, chr4, chr5, chr6, chr7, chr8, chr9, chr10, chr11, chr12, chr13, chr14, chr15, chr16, chr17, chr18, chr19, chr20, chr21, chr22, chrX, chrY, chrM]
##### ERROR   reference contigs = [chrM, chr1, chr2, chr3, chr4, chr5, chr6, chr7, chr8, chr9, chr10, chr11, chr12, chr13, chr14, chr15, chr16, chr17, chr18, chr19, chr20, chr21, chr22, chrX, chrY]

[Sam]

That basically mean that the sorting of your reference and your reads are not the same (Notice that chrM is first in reference whereas it is the last for your reads)

[aj123]

Ok I tried adding the chrM line in the last and now I'm getting:

##### ERROR MESSAGE: Couldn't read file /data/odity/REF_GENOME_hg19/hg19_karyotypic.fa because Found invalid line in index file:chrM    16571   6       50      51

[Sam]

You will have to rebuild the index of the fasta.

And instead of adding new answers, I guess you can just use the add comment function.