After long-read validation, four independent axes of evidence are evaluated and applied as Boolean INFO flags. All four labels can co-occur in any combination.
The variant is supported by both short-read sequencing and tissue-matched PacBio long-read sequencing, with each platform independently exceeding a coverage-scaled read count threshold (Poisson-derived, error rate 0.1%, p < 0.01). Agreement between two sequencing technologies with fundamentally different error profiles — PCR-based Illumina and single-molecule PacBio — provides strong evidence against a technology-specific artifact.
The variant was independently identified by 2 or more distinct variant callers, across all sequencing cores considered together. Callers in different cores count toward this threshold. Agreement between algorithms with different underlying error models and calling strategies argues against a caller-specific artifact. The full set of supporting callers is recorded in the CALLERS INFO field.
The variant passes the per-core read support gate in 2 or more sequencing cores from the same tissue. Each core is evaluated independently; a core contributes to this flag only if it originally called the variant and its read counts meet the coverage-appropriate threshold. Detection across physically distinct tissue sections prepared and sequenced independently argues against a section-specific or library-preparation artifact.
Short-read pileup at this position clears the read-support threshold (p < 1×10⁻⁵, sequencing error rate 0.1%) in 2 or more tissues from the same donor. The IDs of all supporting tissues are listed in the SR_TISSUE_PRESENCE INFO field. A variant present in multiple anatomically distinct tissues is more likely to have arisen early in development and is less likely to be a tissue-preparation artifact compared to a variant detected in only one tissue.