FastaAlternateReferenceMaker is not outputting fasta sequence for multiple samples from a VCF

Question

[anikcropscience]

Hello, I have the coordinates for a gene of interest and I am trying to obtain a consensus sequence from a multi-sample VCF file where eventually I would like to have one fasta sequence for each sample. And then I will perform a multiple sequence alignment. I am using the below command:

gatk FastaAlternateReferenceMaker -R Ref.fasta -O heatshock.fasta -L Scaffold_9__1_contigs__length_70652527:6009958-6011175 -V Multi_sample.vcf 

But the code is only outputting only one fasta sequence and it does not even print the sample name. I have 99 samples in my VCF file.

Can you please tell me what am I doing wrong here? Do I need to provide a vcf for each individual to obtain a sequence for each individual?

Thank you.

Answer 1

[Pierre Lindenbaum]

use --use-iupac-sample https://gatk.broadinstitute.org/hc/en-us/articles/360037594571-FastaAlternateReferenceMaker#--use-iupac-sample

something like:

bcftools query -l Multi_sample.vcf  | while read S
do

gatk FastaAlternateReferenceMaker -R Ref.fasta -O "${S}.fasta" --use-iupac-sample "${S}" -L Scaffold_9__1_contigs__length_70652527:6009958-6011175 -V Multi_sample.vcf 

done

[anikcropscience]

Hi Pierre, Thank you very much. That worked very well. But I have another question. In some of the samples, the fasta output files contain letters like R M Y etc. instead of nucleotide bases. But in other files, the sequences are good.

Can you please tell me why is it happening and how can I correct that? Thanks.