Mitochondrial sequence data analysis

Question

[bandanaschapagain]

Hi All, I am analysing mitochondrial genomic sequence data sets to perform mutation rate analysis among my samples. I performed variant calling by using bcftools and have vcf files. At the time of filtration I need to consider the heteroplasmy state of the mitochondria among the samples. Will keeping the variants supported by low allele depth address the heteroplasmy state?

I am really confused as mitochondria are haploid and heteroplasmic what will be the combination of parameters for variant filteration?

Also can anyone explain me the basic of how genotpye and allele depth related in vcf file?

[bandanaschapagain]

I highly appreciate any help at this point.

Answer 1

[shelkmike]

When I looked for heteroplasmic variants in a mitochondrial genome that had frequency at least 10%, read coverage at least 10x and quality at least 40, I used the following commands:

freebayes -f $reference --ploidy 1 --min-alternate-fraction 0.1 --min-alternate-count 10 --pooled-continuous mapping.sorted.bam > variants_prior_to_the_variant_quality_filtering.vcf

vcffilter -f "QUAL > 40" variants_prior_to_the_variant_quality_filtering.vcf >variants_after_the_variant_quality_filtering.vcf

[bandanaschapagain]

Hi, I may be asking a very basic question here but I am using bcftools with ploidy =1. I have vcf file. I am stuck if or not I can calculate the frequency of alt variant with allele depth. My logic is If AD value for alt = 20 Read depth per sample is 3537 then Alt alle frequency = 20/3537*100 I am not sure if I am in the right direction.