Overview #
Segregation analysis evaluates how a variant tracks with disease within families and remains one of the most direct ways to assess clinical relevance.
In the ACMG/AMP framework, PP1 supports pathogenicity when a variant co-segregates with the disease, while BS4 supports benignity when it fails to segregate as expected.
Although segregation data are rarely available from public databases, SeqSMART combines AI-powered literature mining with a built-in pedigree-based segregation analyzer to help users assess these criteria efficiently and transparently.
1. What Are PP1 and BS4? #
| Criterion | Definition | Evidence Direction |
| PP1 – Co-segregation with Disease in Multiple Affected Family Members | “Co-segregation with disease in multiple affected family members in a gene definitively known to cause the disease.” | Supports pathogenicity when a variant is consistently present in affected family members and absent in unaffected ones, following the expected inheritance pattern. |
| BS4 – Lack of Segregation in Affected Members | “Lack of segregation in affected members of a family.” | Supports benignity when a variant is found in unaffected individuals or absent in affected ones, particularly in fully penetrant diseases. |
The strength of PP1 (Supporting, Moderate, or Strong) increases with the number of informative meioses — independent inheritance events demonstrating consistent co-segregation.
For BS4, confidence increases when clear evidence shows that the variant’s presence and the clinical phenotype are incongruent.
2. SeqSMART’s Two-Tiered Evaluation System #
To maximize coverage and reliability, SeqSMART employs a two-tiered strategy for PP1 and BS4 evaluation:
Tier 1: NLP-Based Evidence Mining #
SeqSMART’s Natural Language Processing (NLP) model automatically scans multiple data sources for references to familial segregation:
- ClinVar records mentioning family studies or segregation notes
- PubMed full-text publications and supplementary materials
- Other genomic resources and variant databases
The NLP engine identifies:
- Descriptions of family-based inheritance studies
- Mentions of zygosity, transmission patterns, or carrier status
- Statements about segregation or non-segregation outcomes
All extracted results are then reviewed by SeqSMART experts to confirm:
- Family structure and relationship accuracy
- Whether the evidence supports PP1 or BS4
- The number and independence of informative meioses
⚠️ Note: Segregation data in public sources are limited and often incomplete. For many variants, PP1 and BS4 cannot be confidently assessed based solely on published information.
Tier 2: Case-Based Segregation via the Pedigree Tool #
SeqSMART’s interactive pedigree builder provides a dynamic, case-based method for evaluating PP1 and BS4 using user-supplied family data.
Step 1: Input Family Genotypes and Phenotypes #
Users can:
- Define family structure and relationships
- Assign each family member’s genotype (e.g., heterozygous, homozygous, wild type)
- Annotate phenotypic status using HPO terms or OMIM diagnoses
Step 2: Automated Inheritance Pattern Detection #
SeqSMART’s segregation analyzer automatically detects whether the variant’s inheritance correlates with the observed phenotype:
- If the variant tracks with affected individuals, PP1 may apply.
- If the variant occurs in unaffected individuals (or is missing in affected ones), BS4 may apply.
The analyzer considers:
- Expected inheritance pattern (dominant, recessive, X-linked, etc.)
- Zygosity consistency
- Penetrance assumptions defined in case settings
Step 3: Dynamic Criterion Assignment #
Once analysis is complete:
- PP1 is marked as Met when significant co-segregation is observed.
- BS4 is marked as Met when non-segregation is evident.
- If data are ambiguous or incomplete, both criteria remain Unassessed.
3. Example #
Autosomal Dominant Case:
- Three affected family members each carry the variant.
- No unaffected members carry the variant.
→ PP1 (Supporting or Moderate) is met depending on the number of informative meioses.
Autosomal Dominant Case (Opposite Pattern):
- Variant present in multiple unaffected relatives.
- Disease is known to be fully penetrant.
→ BS4 (Supporting or Strong) may be applied.
4. Using the Pedigree Tool #
To utilize segregation analysis in SeqSMART:
- Open the Pedigree Builder for the case.
- Add family members and define their relationships.
- Assign variant genotype (from sequencing results or manual entry).
- Tag phenotype data for each member using HPO or OMIM terms.
- Run Segregation Analysis — the system automatically suggests whether PP1 or BS4 is applicable.
Results appear in the variant detail view under Segregation Evidence, along with a visual pedigree diagram for context.
5. Transparency and Expert Control #
- All evidence — whether from NLP mining or user-entered pedigree data — is visible to reviewers.
- Experts may adjust PP1 or BS4 manually if additional evidence is available (e.g., unpublished family data or clinical reports).
- Each change is logged with author identity, timestamp, and justification, ensuring full traceability.
Summary #
SeqSMART integrates both literature-derived and case-based segregation analyses to assess PP1 and BS4 accurately.
By combining NLP-driven data discovery with an AI-powered pedigree analyzer, SeqSMART enables precise, evidence-based evaluation of familial inheritance — even when public data are scarce.
SeqSMART Segregation Principle:
Interpret inheritance intelligently — connect evidence to family context.