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Questions tagged [variant-calling]

The process of discovering variants in individuals starting from sequencing reads. Can be subdivided into germline variant calling and somatic variant calling. Can also be used for the more specific process of finding variants starting with reads aligned to a reference genome (SAM/BAM files).

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pharmacogenomics diplotype calling for CYP2D6 gene from a WES sample

I am currently working on pharmacogenomics diplotype calling for CYP2D6 gene from a WES sample (HG02373) from 1000 genomes, using Aldy4 v4.5, python v 3.11, OS used here in ubuntu linux v22.04, using ...
Neeleshwar Pandey's user avatar
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concatenating windowed VCF files, but quickly?

I am calling variants from WGS for a genetically diverse species which, with my sample size, produces a (bgzipped) VCF of around 500G. I am using a nextflow pipeline to chunk up the genome, call ...
nreid's user avatar
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How do I quantify a specific somatic variant?

I'm working with targeted Illumina sequencing data generated with DNA from diseased and healthy tissue (this is an age-related disease and is not cancer/neoplastic). My hypothesis is that the diseased ...
Nereus's user avatar
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Variants from multiple tools: normalization before or after annotation with VEP?

We want to compare some variant calling tools and their calls on Whole Exome Sequencing data. We will have to normalize the variants that are called (.vcf format) before comparison using for instance ...
Dandelion's user avatar
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bwa mem hangs after a few thousand reads

I am trying to align a bunch of paired sample fastq files using bwa mem. My original command was: ...
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ERROR IN FUNCOTATOR: [SQLITE_IOERR_LOCK] I/O error in the advisory file locking logic (disk I/O error)

I ran the "funcotator" step from the GATK somatic short variant discovery (snvs + indels) pipeline and used the output of filtermutectcalls as input, but ...
Rita Soares's user avatar
1 vote
2 answers
80 views

How to run a GATK Docker Image with local files?

I'm trying to use the HaplotypeCaller from the GATK toolkit but I keep getting an error. I pulled GATK through Docker and am using this command: ...
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which SNP effect software, such as VEP, annovar or snpEff, allows the user to submit their own annotations in gtf format?

I have a use case where I want to annotate the variants from running Oxford Nanopore's Medaka on a very simple reference, a single sequence starting on an ATG and finishing on a stop codon (no introns)...
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INDELS in PLINK files converted to VCF

I want to compare/validate variants called from sequencing data with array (plink format) variant data. I converted the plink files (.bim, .bed, and .fam files) with plink1 to vcf files. ...
Dandelion's user avatar
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Choosing the best number of species for assay

I am doing amplicon sequencing of a virus across many different regions. Lets say I have 20k unique variants of the same virus that I put into my pcr assay and after sequencing and amplifications I am ...
TheNumber23's user avatar
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Feasible to find genetic variations of two samples using RNAseq data?

I have bulk RNAseq data from two strains of mice from Jackson Lab: C57BL/6 and B6.SJL. The former expresses a Ptprc-b allele and the latter expresses a ...
geom_na's user avatar
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Different human reference genomes from NCBI and ENSEMBL, and variant annotation with Variant Effect Predictor (VEP)

Background: I have been working on a human variant calling pipeline, from whole exome reads to variant annotation. For variant annotation I used the Variant Effect predictor (VEP), which is Ensembl ...
Dandelion's user avatar
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Which type of variant caller should I use in a WES normal cell line sample?

I have whole-exome sequencing data of an immortalised non-tumor (normal) cell line that I wish to assess for the presence/absence of APC/Wnt mutations. This is to double check that the cell line is ...
kane9530's user avatar
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Getting VCF file that contain common SNPs from 6 VCF file using isec

I have 6 VCF files, where I would like to obtain the SNPs that are common (by position) in all the 6 files. I have tried this command ...
Mohamed Samir's user avatar
2 votes
1 answer
64 views

WES variant calling with DNBSEQ-T7: technical quality assessment

I recently received whole exome sequencing samples that were sequenced on an MGI sequencing instrument, DNBSEQ-T7. I am interested in somatic variant-calling on the paired tumor-normal samples. As ...
kane9530's user avatar
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Identifying somatic mutations in cell lines

I would like to identify the somatic mutations present in a cell line and characterise the genes that are potentially affected by those mutations. For example, are there oncogenes mutated in a ...
Macintosh's user avatar
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Non-ACGTN alternate alleles in VCF

I was trying to left normalize my vcf file using following command: bcftools norm -f reference.fasta -m -any input.vcf > output.norm.vcf However this gives the ...
Macintosh's user avatar
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variant visualisation of VCF files for 30+ samples

I am relatively new to genome analysis and would like to compare 30+ vcf files, 1 sample per file. After filtering there are about ~20 variants per vcf file. I would like to evaluate to mutational ...
Macintosh's user avatar
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1 answer
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How do I generate a variant list using Illumina reads from a Salmonella genome?

I am planning on performing phylogenetic analysis of Salmonella specimens using WGS data from PulseNet and GenomeTrakr, for the purpose of public health surveillance & to provide context for ...
Erik D's user avatar
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Benchmarking for variant identification using RNA-seq data

I am in need to benchmark the variant identification pipeline which uses RNA seq data alone without any matched-normal. I would like to know the reference dataset (and the pipeline on which the ...
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Compare VCF files

I have two strains, X and Y, and I would like to compare the variants found in both. I have my own parent strain, Z, that was sequenced and assembled to be used as the reference genome. I first ...
rimo's user avatar
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Name this alternative INDEL format

I have recently been working with variants which are represented in a non-standard format. This alternative format uses a period instead of prefixing variants with a reference base. What is this ...
Moss Richardson's user avatar
4 votes
1 answer
124 views

Find SNPs in yeast genomes

I'm a new Bioinformatic scientist working for a yeast genetics company. Objective To create a database of yeast genomes from NCBI and identify SNP variants. In my pipeline FastQC, Trimmomatic, BWA ...
rimo's user avatar
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Difference between pileup and mpileup in VarScan and samtools

I'm going to call variants with VarScan from a pileup files created with samtools. I realized that there are in general two major possibilities to call variants, pileup as well as mpileup. The VarScan ...
Anti's user avatar
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snakemake tutorial stuck in basics-step-5 (variant calling)

I am learning to use snakemake. I am following the tutorial (https://snakemake.readthedocs.io/en/stable/tutorial/tutorial.html). In the section called "Basics", steps 1 to 4 worked well, ...
Sergio.pv's user avatar
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1 answer
257 views

somatic SNV tool for ONT samples

I'm looking for a tool to call somatic single nucleotide variations on reads from an Oxford Nanopore Technologies (ONT) run. I have both tumor and normal samples and have already aligned them to a ...
Walter Gallego Gómez's user avatar
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17 views

How deep variant and others preprocessed insertion and deletion for extracting candidate variant?

I am trying to build a model for variant calling,but I am a bit confused how to pre-process the reads that has insertion and deletion and align them with reference? Should I add something to the ...
Ali Akay's user avatar
1 vote
1 answer
77 views

Reference variant detected as altered one in bam file

I received (from manufacturer) several .bam files and I used four callers (samtools, freebayes, haplotypecaller, deepvariant) to find some sequence variants. In obtained .vcf files, I took a closer ...
Adamm's user avatar
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What is the best tools to find all the somatic mutations?

I am a beginner in this field. I would like to know which is the best way to get all the mutations from a certain sample. At this point, I am considering using GATK (here) and Maftools (here). I don't ...
Scott XU's user avatar
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2 answers
182 views

Detect mutation context in a read of a sam file

After sequencing (Illumina) of some DNA, I generated a sam file through alignment of a fastq file (using Bowtie2). Instead of using variant calling programs, I want to know the specific variances, ...
Annelisa's user avatar
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1 answer
313 views

Range of values for VAF in heterozygous genotype

(VAF = variant allele frequency) This question follows from this one. What is the expected range of values for VAF of mutations with heterozygous genotype (i.e., 0/1 or 0|1)? Is it possible to have 0% ...
gc5's user avatar
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2 votes
0 answers
185 views

Assess ploidy estimation through SNV and CNV results

I want to assess ploidy estimation (i.e., diploid vs tetraploid) in a set of samples for which I have both results from the variant caller (i.e., SNVs) and from the copy number caller. What are the ...
gc5's user avatar
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1 vote
1 answer
658 views

GATK variant filtration using "SelectVariants" and use of JEXL queries

How to do variants selection in some corner cases using GATK and JEXL expressions? I am following the guidelines given in this links for variant selection for some specific cases are not as much ...
everestial's user avatar
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64 views

When can we have duplicated variants in a VCF file, excluding artifacts?

I read these two questions on duplicated VCF variants: 1, 2. However, it is not entirely clear to me when duplicated variants can arise (excluding merging artifacts). I want to understand if ...
gc5's user avatar
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2 votes
2 answers
122 views

Nanopore variant calling

So far I haven't done any variant calling as such. Nanopore I have used for 16s microbiome data. Now My question/doubt so how do I proceed for nano-pore virus sequencing data Steps: I get fast5 files ...
kcm's user avatar
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1 vote
1 answer
188 views

Meaning of order of alleles in VCF GT field for unphased genotypes

Order of alleles in genotype field (on VCFs) can be used to infer haplotypes However, what about unphased genotypes? I have a VCF file where I only have unphased genotypes: ...
gc5's user avatar
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0 votes
1 answer
196 views

Convert VCF file to mpileup.txt

I am working on an iterative analysis that uses orthologous pipelines that require mpileup.txt files as input for a visualization step. This requires me to convert VCF files to mpileup.txt. This ...
Andrew Judell's user avatar
1 vote
2 answers
628 views

How do I call variants using BCFTools without piping mpileup file?

I need to call variants on a large number of reads using both VarScan2 and BCFTools. Both of these variant calling tools use <...
PPrasai's user avatar
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0 answers
36 views

Fastest way to count where two bam files are homozygote reference (inverted variant calling)

Maybe I'm having a brain-freeze so excuse me if this is a waste of everyone's time... I am working with an organism with ~700Mb genome. I have bam files of two individuals that are whole genome ...
AWE's user avatar
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7 votes
4 answers
1k views

Are variant calling files personally identifiable information?

This question is to be read in the context of data protection. It seems to be common understanding that the whole genome sequence of an individual is personally identifiable, non-anonymizable ...
Eekhoorn's user avatar
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2 votes
1 answer
201 views

Consensus sequence from vcf file

I am generating a consensus sequence for SARS-Cov-2. I am visually inspecting every candidate SNP in a pileup. Sometimes I find that there are two consistent variants for the same position, but I can ...
juanjo75es's user avatar
2 votes
1 answer
56 views

Detecting SARS-CoV-2 variants from the mixed virus population

I have a fastq file from Nanopore sequencing data that contains reads from both the UK and South Africa variants of SARS-CoV-2. The variants are identified by three key mutations in the receptor-...
L R Joshi's user avatar
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1 vote
2 answers
85 views

Solutions to reference bias issue

I've recently learned about the reference bias issue - inability to properly map NGS reads in certain genomic regions caused by the fact that our standard genomic references are linear and do not ...
Igor's user avatar
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3 votes
2 answers
62 views

Calling for one specific SNP from multiple sequencing runs

Just learning the basics of bioinformatics and bash shell scripting. Is there a correct way to search for a specific allele with one or even multiple sequencing runs? I am searching for prevalence of ...
S.J.A's user avatar
  • 31
2 votes
2 answers
2k views

Can an alternate allele be more common than a reference allele?

I'm trying to analyze data from a genotype-phenotype association study. The genotype data is stored in csv files that record for each variant site and each subject, the alleles that the subject has at ...
J.D.'s user avatar
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1 vote
0 answers
138 views

SnpEff not correctly annotating multiple-nucleotide polymorphisms

I have some SARS-CoV-2 sequencing data that I'm trying to annotate with SnpEff, however SnpEff doesn't appear to be recognizing multiple adjacent SNPs within the same codon and correctly calling them ...
Set's user avatar
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1 vote
1 answer
172 views

Why is there no GT-entry in my .vcf file from bacteria

I mapped raw reads obtained from an E. coli genome (.fastq file) to my reference genome. Next, I exported the .vcf (variant calling file), which looks something like the header example shown below (...
David Streuli's user avatar
1 vote
1 answer
290 views

genotyping or variant calling in R

Are there any good options for calling variants or even just a pileup from R? R is not ideal for this, but I'd like to integrate with other functions. I found deepSNV::bam2R which roughly does what I ...
burger's user avatar
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0 votes
1 answer
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Supplementary aligments in VAF

This question has also been asked on Biostars I have a doubt, are supplementary alignment usually considered when the variant allele frequency is calculated? Thanks a lot. In my case I have some ...
Denise Lavezzari's user avatar
2 votes
1 answer
158 views

Are there open-sourced graph-based variant callers?

I would like to compare GATK with graph-based variant callers. I have seen the Fast and accurate genomic analyses using genome graphs paper from SevenBridges and the paragraph tool by Illumina, though ...
0x90's user avatar
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