147: Full-length ABO Haplotype Sequencing and Variant Resolution

Ying Y et al., Clinical Chemistry 71:4 (2025) 510–519 - This episode reviews a Clinical Chemistry study that developed an improved one-step ultra-long-range PCR with PCR suppression primers and PacBio SMRT long-read sequencing to obtain 26.1 kb full-length ABO haplotypes from the 5′ UTR to the 3′ UTR, enabling comprehensive allele annotation and resolution of complex ABO variants. Key terms: ABO, haplotype sequencing, long-read sequencing, structural variants, 3' UTR.

Study Highlights:
The authors amplified a 26.1 kb full-length ABO amplicon using an improved one-step ultra-long-range PCR with PCR suppression primers and generated phased haplotypes by PacBio SMRT sequencing. They analyzed 158 haplotypes from 79 blood donors and 47 variant specimens, identifying coding, intronic, promoter, UTR, and structural variants. The work revealed allele-specific intron 1 (TA)n VNTR patterns, detailed 5′ and 3′ UTR motif/unit structures, and resolved large SVs including a 7337 bp deletion in an erythroid enhancer, recombinants, and chimeras. The method outperformed PCR-SBT and short-read NGS for phasing and SV detection but requires high-quality DNA and limited UTR/intronic validation by secondary methods.

Conclusion:
Full-length ABO haplotype sequencing spanning 5′ to 3′ UTR fills reference gaps, refines allele sequence patterns, and improves detection and resolution of structural and regulatory variants relevant to transfusion and transplantation; practical use is powerful but constrained by DNA quality and limited orthogonal validation for some noncoding variants.

Music:
Enjoy the music based on this article at the end of the episode.

Article title:
Comprehensive Annotation of Complete ABO Alleles and Resolution of ABO Variants by an Improved Full-Length ABO Haplotype Sequencing

First author:
Ying Y

Journal:
Clinical Chemistry 71:4 (2025) 510–519

DOI:
10.1093/clinchem/hvaf015

Reference:
Ying Y, Zhang J, Hong X, Yuan W, Ma K, Huang X, Xu X, Zhu F. Comprehensive Annotation of Complete ABO Alleles and Resolution of ABO Variants by an Improved Full-Length ABO Haplotype Sequencing. Clinical Chemistry. 2025;71(4):510-519. doi:10.1093/clinchem/hvaf015

License:
This episode is based on an open-access article published under the Creative Commons Attribution 4.0 International License (CC BY 4.0) – https://creativecommons.org/licenses/by/4.0/

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Episode link: https://basebybase.com/episodes/comprehensive-annotation-of-complete-abo-alleles-and-resolution-of-abo-variants

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-09-24.

QC Scope:
- article metadata and core scientific claims from the narration
- excludes analogies, intro/outro, and music
- transcript coverage: Audited the transcript sections describing (1) end-to-end ABO haplotype sequencing from 5' UTR to 3' UTR, (2) ultra-long-range PCR with PCR suppression, (3) PacBio SMRT sequencing and haplotype phasing, (4) haplotype patterning and structural variations including deletions, recombination, and chimeras, and (5) clinical
- transcript topics: Full-length ABO haplotype sequencing (5' UTR to 3' UTR); Ultra-long-range PCR with PCR suppression; PacBio SMRT sequencing and CCS/pbmm2 analysis; Intron 1 TA VNTR and 5' UTR minisatellites; 3' UTR unit mapping (units 1–14) and NG_006669.2 comparison; Structural variations: deletions, recombination, chimeras

QC Summary:
- factual score: 10/10
- metadata score: 10/10
- supported core claims: 7
- claims flagged for review: 0
- metadata checks passed: 4
- metadata issues found: 0

Metadata Audited:
- article_doi
- article_title
- article_journal
- license

Factual Items Audited:
- 26.1 kb full-length ABO haplotype amplicon with no splicing
- 158 haplotypes from 79 donors and 47 ABO variants
- Dominant alleles A1.01, A1.02, B.01, O.01.01, O.01.02; two rare alleles A2.05 and O.01.71
- TA VNTR in intron 1; median TA repeats: 21 for A1.02 and B.01; 13 for O alleles
- 5' UTR minisatellite motifs and 3' UTR units 1–14; NG_006669.2 lacks units 11–13
- 7337 bp deletion in an erythroid enhancer associated with Bel phenotype

QC result: Pass.