---
title: Why can two callers show different VAFs for the same variant?
description: A guide to understanding why Mutect2 and Freebayes can report different Variant Allele Frequencies (VAFs) for the same somatic variant. Learn about read realignment and how to interpret conflicting results.
---
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# Why can two callers show different VAFs for the same variant?
## When two variant callers disagree: discover the technical reasons for VAF differences, and learn how to interpret these discrepancies in your somatic analysis.
Variant callers sometimes detect and normalize the same underlying event differently. This can lead to mismatched coordinates, diverging allele fractions, and inconsistent downstream annotations.
- **Repeat motifs and indel representation**: In regions with motif repeats, callers can legitimately place indel breakpoints at slightly different positions. This shifts the reported coordinates and prevents automatic merging into a single entry.
- **Normalization choices**: Freebayes and GRIDSS can normalize the same event differently. Those differences are valid but not identical in VCF coordinates.
---
### In SeqOne's Somatic analyses: why can the VAF differ between Mutect2 and Freebayes?
- **Mutect2** realigns reads during calling. This on-the-fly realignment can change which reads support the variant and therefore the VAF.
- **Freebayes** analyzes the raw alignment without runtime read realignment. Its VAF will more closely reflect what you see in IGV when viewing the BAM, which is also pre-realignment.
Implications
- It is normal for VAFs to diverge, especially for indels and complex events.
- IGV’s read counts typically align better with Freebayes than with Mutect2 because IGV shows the original BAM, not the Mutect2 realigned view.
---
### In SeqOne's Somatic analyses: how to visualize VAF discrepancies in the Variant Viewer?
- When VAFs conflict, both Mutect2 and Freebayes values are shown side-by-side so you can assess the difference.
- Data-type aware prioritization in the Variant table:
- Capture workflows: Freebayes information is prioritized.
- Amplicon workflows: Mutect2 information is prioritized.
- A visual warning when a notable VAF discrepancy exists in Amplicon analyses, to draw attention during review.
---
### How to interpret discording VAFs?
- 🧬 Consider biology and context. For indels and repetitive regions, expect larger VAF discrepancies.
- 🔍 Cross-check with IGV to understand evidence in the raw BAM.
- 🤝 Use both callers’ outputs as complementary evidence rather than picking a single "winner".
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