Recalibrated output (BQSR)

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Maggie Sudo Pui San

• June 04, 2021 01:44

Hi, I have been using GATK version 4.2 for my work. The recalibration plot seemed to be acceptable ... However my cycle covariate plots looked a bit off. They have low log10 (observations) values on the negative scale (-1 to -100++) of the x-axis... Even in the other plots, the log10 value seemed to be very low/ faint too. Is that normal?

The commands that I used from variant filtration to Analyse Covariate are listed below:

VariantFiltration 

./gatk --java-options "-Xmx128g" VariantFiltration -R reference -V snps.vcf -O filtered_snps.vcf -filter-name "QD_filter" -filter "QD < 2.0" -filter-name "FS_filter" -filter "FS > 60.0" -filter-name "MQ_filter" -filter "MQ < 30.0" -filter-name "SOR_filter" -filter "SOR > 4.0" -filter-name "MQRankSum_filter" -filter "MQRankSum < -12.5" -filter-name "ReadPosRankSum_filter" -filter "ReadPosRankSum < -8.0"

./gatk --java-options "-Xmx128g" VariantFiltration -R reference -V indels.vcf -O filtered_indels.vcf -filter-name "QD_filter" -filter "QD < 2.0" -filter-name "FS_filter" -filter "FS > 200.0" -filter-name "SOR_filter" -filter "SOR > 10.0"

Select Variants

./gatk --java-options "-Xmx128g" SelectVariants --exclude-filtered -V filtered_snps.vcf -O bqsr_snps.vcf

./gatk --java-options "-Xmx128g" SelectVariants --exclude-filtered -V filtered_indels.vcf -O bqsr_indels.vcf

BaseRecalibrator

./gatk --java-options "-Xmx128g" BaseRecalibrator -R reference -I Sample.bam -known-sites bqsr_snps.vcf --known-sites bqsr_indels.vcf -O data.table

Apply BQSR

./gatk --java-options "-Xmx128g" ApplyBQSR -R reference -I Sample.bam -bqsr data.table -O datarecal_reads.bam

Base Recalibrator

./gatk --java-options "-Xmx128g" BaseRecalibrator -R reference -I datarecal_reads.bam -known-sites bqsr_snps.vcf --known-sites bqsr_indels.vcf -O post_data.table

AnalyzeCovariates

./gatk --java-options "-Xmx128g" AnalyzeCovariates -before data.table -after post_data.table -plots recalibration_plots.pdf

 

Thank you in advance!

 

 

 

 

 

 

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  Maggie Sudo Pui San

  • June 04, 2021 01:52

  I forgot to mention what I was trying to do in the previous post.

  I am performing variant calling for my whole genome sequences. The variants will be used to assess their phylogenetic relationship.

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  Bhanu Gandham

  • June 04, 2021 22:00
  • Edited

  Hi Maggie, 

  These values look normal to me. Take a look at this article for detailed documentation: https://gatk.broadinstitute.org/hc/en-us/articles/360035890531-Base-Quality-Score-Recalibration-BQSR-

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  Maggie Sudo Pui San

  • June 07, 2021 04:59

  Hi Bhanu, 

   

  Thank you! I'm relieved to hear that. I will check out the article for detailed documentation. Thanks. 

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