mutect2 results

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Liroz Hafouta

• December 06, 2024 08:15

Hi all, I'll be happy to get your help.

Im doing a process of variant calling through Mutect2 tool (in normal-tumor mode) and VEP, to analyze somatic variants appearing in ovary tumors. Im havig a hard time to understand all INFO column meaning and have several questions. 

1. is there a guide you wrote about the meaning of all VCF results and columns? I see no publications so it is difficult to deeply understand the way you calculated everything.

2. for some reason the NALOD index is positive in all variants Im dealing with, which I assume is not good enough to reflect true results instead of artifacts. I wonder how if you can explain to me what does the NALOD reffers to, and if it is still OK if it is positive (values are between 0-2 and a little more) to relay on the results that they are not an artifact?

3. How can I validate the results I receive to know if the variants are truely correct? It is a clinical study and therefore it is crucial for us to be accurate. 

thank you in advance,

Liroz

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• 

  Gökalp Çelik

  • December 09, 2024 08:25
  • Edited

  Hi Liroz Hafouta

  NALOD annotation is better explained here in the document in our github repository. More of our annotations can be found in this document as well.

  https://github.com/broadinstitute/gatk/blob/master/docs/mutect/mutect.pdf 

  

  In summary Mutect2 applies a somatic model to the normal sample to detect the presence of possible artifacts that is present in the tumor sample. This value is a P value calculated by the somatic likelihoods and returned as a negative log 10 value for simplicity. This value is then used by FilterMutectCalls tool to apply somatic filters onto the raw Mutect2 VCF. The filter applied for the NALOD can be adjusted by the parameter 

  --normal-p-value-threshold <Double>
                                P value threshold for normal artifact filter  Default value: 0.001.

  in the FilterMutectCalls tool. So any sites that have NALOD value 3 or more will get filtered with Normal artifact filter. If NALOD value is less than 3 then p value gets less significant for the filtering based on the default value. If you wish to set this default value as 0.05 that means any artifact in normal with NALOD 1.3 or more will get filtered (you may calculate this conversion by log10(pvalue)*-1).

  As for the clinical study the you need to confirm the presence of variants with orthogonal methods like sanger sequencing or qPCR, resequencing etc... 

  I hope this helps. 

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