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How to generate variant calls with bcftools

This recipe demonstrates variant call generation using bcftools and simulated data.

1 result • updated 5.1 years ago by Istvan Albert

This recipe demonstrates variant call generation using bcftools and simulated data.

When using simulations the variants will be known and we can use the process to evaluate the accuracy of the variant caller.

The installation commands in the book do not include dwgsim. It will need to be installed separately with:

conda install dwgsim

For more information see the

Bioinformatics Data Analysis online course.
Biostar Handbook for system setup and other information.

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# Stop on errors.
set -uex

# Reference accession numbers.
ACC=AF086833

# Create the directory for reference file.
mkdir -p refs

# The name of the reference.
REF=refs/${ACC}.fa

# The name of the BAM file
BAM=align.bam

# Obtain the reference genome.
efetch -db=nuccore -format=fasta -id=AF086833 > $REF

# Create a bwa index for the reference.
bwa index $REF 1>>log.txt 2>> log.txt

# Create a samtools index for the reference.
samtools faidx $REF

# Simulate reads from the reference file.
# Generates the variant file called simulated.mutations.vcf
dwgsim $REF simulated 2>> log.txt

# This is the read naming generated by dwgsim.
R1=simulated.bwa.read1.fastq
R2=simulated.bwa.read2.fastq

# Generate the alignment from the data simulated above.
bwa mem $REF $R1 $R2 2>> log.txt | samtools sort > $BAM 2>> log.txt

# Index the alignment
samtools index $BAM

# Compute the genotypes from the alignment file with pileup.
bcftools mpileup -B -Ovu -f $REF $BAM > genotypes.vcf

# Call the variants from the genotypes.
bcftools call -vc -Ov genotypes.vcf > observed-mutations.vcf

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[acc]
label = "Genome accession number"
display = "TEXTBOX"
value = "AF086833"
regex = "^\\w{1,20}$"
help = "Must be an NCBI accession number"

[N]
label = "The number of reads"
display = "INTEGER"
value = 100000
range = [ 1, 1000000,]
help = "How many simulated read pairs to generate."

[settings]
id = 68
recipe_uid = "recipe-variants-bcftools"
uid = "recipe-variants-bcftools"
name = "How to generate variant calls with bcftools"
template = "How_to_generate_variant_calls_with_bcftools_bio-data-analysis_68.sh"
image = "How_to_generate_variant_calls_with_bcftools_bio-data-analysis_68.png"
project_uid = "2a988f7f"
url = "http://localhost8000"
help = "This recipe demonstrates variant call generation using bcftools and simulated data. \n\nWhen using simulations the variants will be known and we can use the process to evaluate the accuracy of the variant caller.\n\nThe installation commands in the book do not include `dwgsim`. It will need to be installed separately with:\n\n    conda install dwgsim\n\nFor more information see the\n\n* [Bioinformatics Data Analysis][link]  online course.\n* [Biostar Handbook][book] for system setup and other information.\n\n[link]: https://www.biostarhandbook.com/edu/course/4/\n[book]: https://www.biostarhandbook.com"

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