Why Does Mpileup Skip some variants ?
when I make variant call with bcftools mpileup and call , i can't get some variants,especially those have the same number of Ref and Mutant reads!!!
My command is :
bcftools mpileup -Ou -f ref my.sorted.bam | bcftools call --ploidy 1 -mv -Ov -o my.vcf
I can't find this variant in my vcf :
T (Ref: with depth=94) ---> C (Variant: with depth=94)
This variant exist in IGV but i can't find it in my vcf.
Your question is severely lacking in details. Please show all commands that you have used, and also describe your experimental setup. From my experience, mpileup has greater sensitivity over other variant callers for germline variants.
my command is :
bcftools mpileup -Ou -f ref my.sorted.bam | bcftools call --ploidy 1 -mv -Ov -o my.vcf
I can't find this variant in my vcf :
T (Ref: with depth=94) ---> C (Variant: with depth=94)
This variant exist in IGV but i can't find it in my vcf.
I see.... why --ploidy 1 ?
Great; so, you sequenced a mixed population of bacteria and expect that some of these carry a mutation?
I see no further details. If you sequenced a mixed population of Mycobacterium spp. where you expect 50% of the population to be wild-type and the other [50%] to have a mutation, and are now telling BCFtools mpileup to expect ploidy of 1, then BCFtools will struggle to get the result that you want. Try the other option of Freebayes.
My problem is:
In this position : ====> T (Ref: with depth=94) ---> C (Variant: with depth=94)
T and C have the same depth of coverage , so bcftools don't call this variant.
I need to tell bcftool to call variant when there is the same depth of coverage in the reference and in the mutant allele. BUT I DON'T KNOW HOW TO DO IT
ploidy = 1, BCFtools will struggle to call this variant for you unless you alter other parameters.Good luck with it.
Because i don't have more detail .
FreeBayes is good but it dosen't give me DP4 in vcf.
I have the same problem without ploidy 1 with bcftools!
Do you have any idea how can i force DP4 to be present in vcf ? what parameters should i give to FreeBayes !
When i check DP4 , i get this error : Error: the tag "DP4" is not defined in the VCF header
Thank you!
Please explain in full detail your experimental setup (see my other comments)
You may instead try Freebayes.
Thanks!
My problem is:
In this position : ====> T (Ref: with depth=94) ---> C (Variant: with depth=94)
T and C have the same depth of coverage , so bcftools don't call this variant.
I need to tell bcftool to call variant when there is the same depth of coverage in the reference and in the mutant allele. BUT I DON'T KNOW HOW TO DO IT
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Hello sami!
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