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- Introduced a comprehensive RNA velocity analysis pipeline using scVelo, including data loading, preprocessing, velocity estimation, and visualization. - Added a script for running RNA velocity analysis with customizable parameters and output options. - Created detailed documentation for IQ-TREE 2 phylogenetic inference, covering command syntax, model selection, bootstrapping methods, and output interpretation. - Included references for velocity models and their mathematical framework, along with a comparison of different models. - Enhanced the scVelo skill documentation with installation instructions, use cases, and best practices for RNA velocity analysis.
86 lines
3.5 KiB
Markdown
86 lines
3.5 KiB
Markdown
# gnomAD Variant Interpretation Guide
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## Allele Frequency Thresholds for Disease Interpretation
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### ACMG/AMP Criteria
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| Criterion | AF threshold | Classification |
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|-----------|-------------|----------------|
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| BA1 | > 0.05 (5%) | Benign Stand-Alone |
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| BS1 | > disease prevalence | Benign Supporting |
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| PM2_Supporting | < 0.0001 (0.01%) for dominant; absent for recessive | Pathogenic Moderate → Supporting |
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**Notes:**
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- BA1 applies to most conditions; exceptions include autosomal dominant with high penetrance (e.g., LDLR for FH: BA1 threshold is ~0.1%)
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- BS1 requires knowing disease prevalence; for rare diseases (1:10,000), BS1 if AF > 0.01%
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- Homozygous counts (`ac_hom`) matter for recessive diseases
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### Practical Thresholds
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| Inheritance | Suggested max AF |
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|-------------|-----------------|
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| Autosomal Dominant (high penetrance) | < 0.001 (0.1%) |
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| Autosomal Dominant (reduced penetrance) | < 0.01 (1%) |
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| Autosomal Recessive | < 0.01 (1%) |
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| X-linked recessive | < 0.001 in females |
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## Absence in gnomAD
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A variant **absent in gnomAD** (ac = 0) is evidence of rarity, but interpret carefully:
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- gnomAD does not capture all rare variants (sequencing depth, coverage, calling thresholds)
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- A variant absent in 730K exomes is very strong evidence of rarity for PM2
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- Check coverage at the position: if < 10x, absence is less informative
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## Loss-of-Function Variant Assessment
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### LOFTEE Classification (lof field)
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- **HC (High Confidence):** Predicted to truncate functional protein
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- Stop-gained, splice site (±1,2), frameshift variants
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- Passes all LOFTEE quality filters
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- **LC (Low Confidence):** LoF annotation with quality concerns
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- Check `lof_flags` for specific reason
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- May still be pathogenic — requires manual review
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### Common lof_flags
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| Flag | Meaning |
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|------|---------|
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| `NAGNAG_SITE` | Splice site may be rescued by nearby alternative site |
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| `NON_CANONICAL_SPLICE_SITE` | Not a canonical splice donor/acceptor |
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| `PHYLOCSF_WEAK` | Weak phylogenetic conservation signal |
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| `SMALL_INTRON` | Intron too small to affect splicing |
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| `SINGLE_EXON` | Single-exon gene (no splicing) |
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| `LAST_EXON` | In last exon (NMD may not apply) |
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## Homozygous Observations
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The `ac_hom` field counts homozygous (or hemizygous in males for chrX) observations.
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**For recessive diseases:**
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- If a variant is observed homozygous in healthy individuals in gnomAD, it is strong evidence against pathogenicity (BS2 criterion)
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- Even a single homozygous observation can be informative
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## Coverage at Position
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Always check that gnomAD has adequate coverage at the variant position before concluding absence is meaningful. The gnomAD browser shows coverage tracks, and coverage data can be downloaded from:
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- https://gnomad.broadinstitute.org/downloads#v4-coverage
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## In Silico Predictor Scores
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| Predictor | Score Range | Pathogenic Threshold |
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|-----------|-------------|---------------------|
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| CADD PHRED | 0–99 | > 20 deleterious; > 30 highly deleterious |
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| REVEL | 0–1 | > 0.75 likely pathogenic (for missense) |
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| SpliceAI max_ds | 0–1 | > 0.5 likely splice-altering |
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| SIFT | 0–1 | < 0.05 deleterious |
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| PolyPhen-2 | 0–1 | > 0.909 probably damaging |
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## Ancestry-Specific Considerations
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- A variant rare overall may be a common founder variant in a specific population
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- Always check all ancestry-specific AFs, not just the total
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- Finnish and Ashkenazi Jewish populations have high rates of founder variants
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- Report ancestry-specific frequencies when relevant to patient ancestry
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