filtering a VCF file based on genotype

Question

[Bogdan]

Dear all, I would appreciate having a piece of advice :

how shall I filter a VCF file in order to exclude the GERMLINE and TUMOR samples that have a GENOTYPE of "./".

in R/BioC, for example, if we aim to see the GENOTYPES in the vcf file, we receive the following messages :

> geno(vcf)$GT[,"TUMOR"]
Error in geno(vcf)$GT[, "TUMOR"] : subscript out of bounds
> geno(vcf)$GT[,"NORMAL"]
Error in geno(vcf)$GT[, "NORMAL"] : subscript out of bounds

thank you very much !

[Ram]

What tools are you using? "R/BioC" have multiple tools. Can you please give us the exact commands/scripts you're using?

[Bogdan]

Dear gentlemen, thank you for your replies and help. Here I am posting the R code that I am using to do the filtering of a VCF file that is generated by DELLY and contains INVERSIONs only .. Any suggestions on how to post/link the input VCF file would be welcome. Thank you very much !

library(devtools)
library(stringr)
library(dplyr)
library(ggplot2)

library(VCFWrenchR)
library(VariantAnnotation)
library(VariantTools)

library(GenomicRanges)
library(GenomicFeatures)
library(rtracklayer)
library(org.Hs.eg.db)

library(TxDb.Hsapiens.UCSC.hg38.knownGene)
library(StructuralVariantAnnotation) 


vcf <- readVcf("vcf.INV3.vcf", 
                genome="hg38")

name <- "vcf.INV3.vcf"

TUMOR  <-  samples(header(vcf))[1] 
NORMAL <-  samples(header(vcf))[2]


DEPTH_threshold <- 8
FRACTION_threshold <- 0.2 



PASS_IMPRECISE_filters = function(x) { DV_germline <- ( geno(vcf)$DV[,NORMAL] < 1 )
                                       RV_germline <- ( geno(vcf)$RV[,NORMAL] < 1 )

                                       DV_tumor <- geno(vcf)$DV[,TUMOR]
                                       RV_tumor <- geno(vcf)$RV[,TUMOR]  

                                       DR_tumor <- geno(vcf)$DR[,TUMOR]
                                       RR_tumor <- geno(vcf)$RR[,TUMOR] 

                                       AD <-  (DV_tumor > DEPTH_threshold)      
                                       AF <- ((DV_tumor / (DV_tumor + DR_tumor)) > FRACTION_threshold)

                                       DV_germline &
                                       RV_germline & 
                                       AD &
                                       AF & 
                                       (filt(vcf) == "PASS") & 
                                       (info(vcf)$IMPRECISE) 
                                      }

vcf_PASS_IMPRECISE_FILTERS <- vcf[PASS_IMPRECISE_filters(vcf)]

writeVcf(vcf_PASS_IMPRECISE_FILTERS,
         file=paste(name, ".filter.PASS.IMPRECISE.vcf", sep=""))



PASS_PRECISE_filters = function(x) {   DV_germline <- ( geno(vcf)$DV[,NORMAL] < 1 )
                                       RV_germline <- ( geno(vcf)$RV[,NORMAL] < 1 )

                                       DV_tumor <- geno(vcf)$DV[,TUMOR]
                                       RV_tumor <- geno(vcf)$RV[,TUMOR]  

                                       DR_tumor <- geno(vcf)$DR[,TUMOR]
                                       RR_tumor <- geno(vcf)$RR[,TUMOR] 

                                       AD <- ((DV_tumor + RV_tumor) > DEPTH_threshold)      
                                       AF <- ((RV_tumor / (RV_tumor + RR_tumor)) > FRACTION_threshold) 

                                       DV_germline &
                                       RV_germline & 
                                       AD &
                                       AF & 
                                       (filt(vcf) == "PASS") & 
                                       (info(vcf)$PRECISE) 
                                    }


vcf_PASS_PRECISE_FILTERS <- vcf[PASS_PRECISE_filters(vcf)]

writeVcf(vcf_PASS_PRECISE_FILTERS, 
         file=paste(name, ".filter.PASS.PRECISE.vcf", sep=""))


vcf_PASS_COMBINED_FILTERS <- rbind(vcf_PASS_IMPRECISE_FILTERS, vcf_PASS_PRECISE_FILTERS) 

writeVcf(vcf_PASS_COMBINED_FILTERS, 
         file=paste(name, ".filter.PASS.PRECISE.and.IMPRECISE.and.AD.and.AF.vcf", sep=""))

[ATpoint]

Can you give some background on how the VCF was generated?

[cpad0112]

Please post the program you are using and the sample vcf file.

[Kevin Blighe]

I will add to the comments from the 3 other great people. R would not be my 'go to' environment for filtering a VCF. To do what you want, just use bcftools view with the following parameter:

-m/M, --min-alleles/--max-alleles <int>     minimum/maximum number of alleles listed in REF and ALT (e.g. -m2 -M2 for biallelic sites)

Answer 1

[Jorge Amigo]

from your question I understand that you have 1 single vcf file with 2 samples (germline and tumor) in it, and that you want to filter out all variants where 1 or the 2 samples have an empty genotype (./.)

if this is the case, a simple grep should be enough:

grep -vP '\t\./\.' file.vcf > file.non-empty-gt.vcf

[Bogdan]

Thank you Jorge. Yes, at the end, I have done it with BCFTOOlS:

bcftools view --genotype ^miss $FILE  > "${FILE%.vcf}.fGT.vcf"

I was hoping to do everything in R, as it would be easier to assemble the entire pipeline in R (than to assemble all the scripts I have in a master bash shell .sh script ) .

[Ram]

Unfortunately, R is not the best for all tasks. You're better off using an automation/pipelining tool that is built for interoperability. People recommend snakemake, but I am not sure how well it would fit your use case, as I'm not conversant with snakemake. For the moment, you can either use system() calls from within R or Rscript/R commands within a shell script.

[Bogdan]

Thank you Ram for your help. Yes, i've used the "Rscript" calls from a shell script.

[Ram]

Please use the formatting bar (especially the code option) to present your post better. I've done it for you this time.