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Wen L et al., Trends in Genetics - This review surveys the integration of single-molecule long-read sequencing (SMS) with single-cell genomics, epigenomics and transcriptomics, describing platforms, methods and the new biological 'dark matter' now accessible at single-cell resolution. Key terms: single-cell sequencing, long-read sequencing, alternative splicing, structural variation, transposable elements.

Study Highlights:
SMS platforms (PacBio HiFi and Oxford Nanopore) have improved accuracy and enable direct full-length cDNA sequencing to resolve alternative splicing and isoforms in individual cells. Long reads substantially improve detection of repetitive elements, structural variants, ecDNAs and enable chromosome-scale haplotype phasing from small cell numbers. SMS-based single-cell epigenome methods capture allele-specific DNA methylation, histone marks, TF binding and higher-order 3D chromatin interactions, including within repetitive regions. New single-cell long-read methods (for transcriptome, genome and multiple epigenomic layers) are expanding capabilities but remain constrained by capture efficiency, cost and amplification artifacts.

Conclusion:
SMS-based long-read single-cell omics opens access to genomic and epigenomic 'dark matter'—full-length isoforms, repeats, SVs and allele-specific regulation—and points toward future single-cell de novo assemblies, multi-omics and spatial long-read applications.

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Article title:
Single-cell omics sequencing technologies: the long-read generation

First author:
Wen L

Journal:
Trends in Genetics

DOI:
10.1016/j.tig.2025.07.012

Reference:
Wen L. and Tang F. Single-cell omics sequencing technologies: the long-read generation. Trends in Genetics. 2025. https://doi.org/10.1016/j.tig.2025.07.012

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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On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/the-longread-leap-in-singlecell-omics

Bleidorn C et al., Trends in Genetics - This episode examines a Trends in Genetics review arguing that short-read shotgun sequencing and genome skimming remain powerful, cost-effective tools for biodiversity research. The authors highlight applications from species identification and biomass estimation to phylogenomics and museum-based genomics, and discuss how short reads complement long-read initiatives to meet Global Biodiversity Framework goals. Key terms: short-read sequencing, genome skimming, biodiversity monitoring, museum genomics, phylogenomics.

Study Highlights:
Emerging short-read platforms are lowering costs and increasing throughput, making large-scale sequencing more accessible. Genome skimming recovers organellar genomes and phylogenetically informative nuclear markers from low-coverage data, and assembly-free methods expand phylogenetic analyses to degraded or low-coverage samples. Short-read data enable accurate estimation of genome size and repeat content from skimmed datasets, and museum collections offer a rich source to fill taxonomic and genomic gaps for biodiversity monitoring. Together, these approaches support scalable monitoring and reference database building aligned with the GBF.

Conclusion:
Short-read shotgun sequencing, especially genome skimming and assembly-free analyses, remains a practical and scalable approach to generate genomic resources across taxa, complement long-read reference genomes, and accelerate biodiversity monitoring, phylogenomics, and conservation efforts under the Global Biodiversity Framework.

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Article title:
The untapped potential of short-read sequencing in biodiversity research

First author:
Bleidorn C

Journal:
Trends in Genetics

DOI:
10.1016/j.tig.2025.09.001

Reference:
Bleidorn C, Sandberg F, Martin S, Vogler AP, Podsiadlowski L. The untapped potential of short-read sequencing in biodiversity research. Trends in Genetics. 2025. https://doi.org/10.1016/j.tig.2025.09.001

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/

Support:
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Episode link: https://basebybase.com/episodes/the-untapped-potential-of-shortread-sequencing-in-biodiversity-research

Aston AN et al., Trends in Genetics - This opinion article reviews how short sequence interruptions within expanded tandem repeats alter somatic instability and clinical outcomes across multiple repeat expansion diseases, and proposes synthesis-dependent microhomology-mediated end joining (SD-MMEJ) as a unifying mechanism to explain the gain, loss, and complexity of interruptions. Key terms: repeat expansion, interruptions, SD-MMEJ, DNA repair, Huntington's disease.

Study Highlights:
Interruptions within expanded tandem repeats modulate somatic expansion and clinical severity and are observed to be dynamic, recurrent, locus-specific, and enriched near repeat–flank boundaries. The authors synthesize literature and sequencing observations to show interruptions can arise or be lost within a single generation and can form highly complex alleles. They propose SD-MMEJ, a Polθ-dependent alternative end-joining pathway, as a mechanism that produces insertions, deletions and repeating patterns via priming, synthesis and microhomology annealing after double-strand breaks. The model makes testable predictions about the role of flanking sequence structure, the need for double-strand breaks, and implications for diagnostics and therapeutic manipulation of interruptions.

Conclusion:
SD-MMEJ is presented as a plausible, testable mechanism that can account for most documented interruption dynamics; validating this model could improve interpretation of long-read sequencing, illuminate links between repair pathways and repeat instability, and suggest routes for therapeutic intervention.

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Article title:
Interruptions impact clinical features of repeat expansion diseases, but how are they gained and lost?

First author:
Aston AN

Journal:
Trends in Genetics

DOI:
10.1016/j.tig.2025.07.005

Reference:
Aston AN, Dion V. Interruptions impact clinical features of repeat expansion diseases, but how are they gained and lost? Trends in Genetics (2025). https://doi.org/10.1016/j.tig.2025.07.005

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/

Support:
Base by Base is independent and ad-free — no sponsors, no paywall. If an episode was worth your time, chip in and keep the papers audited and the original songs coming:
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Episode link: https://basebybase.com/episodes/interruptions-in-repeat-expansion-diseases-how-are-they-gained-and-lost

Villalba A et al., Trends in Genetics - A concise exploration of how synthetic DNA could enable engineered haploid gametes, why that possibility intensifies the philosophical non-identity problem, and what this means for reproductive autonomy, disease prevention, and concerns about eugenics in future reproduction. Key terms: synthetic gametes, non-identity problem, reproductive ethics, synDNA, eugenics concerns.

Study Highlights:
The paper argues that advances in synthetic DNA may make de novo construction of haploid gamete genomes feasible, enabling precise control over inherited variants. This capability raises the non-identity problem: creating synthetic gametes may change which individual comes into existence, complicating person-affecting harm assessments. Synthetic gametes could reduce transmission of serious heritable diseases without directly benefiting any specific future person, shifting ethical focus toward impersonal wellbeing and reproductive autonomy. The authors contrast manufacturing gametes with embryo editing and discuss worries about eugenics while emphasizing differences from historical coercive practices.

Conclusion:
Synthetic gamete technology could expand reproductive options and reduce heritable disease but requires renewed ethical frameworks centered on impersonal wellbeing, autonomy, and careful governance.

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Article title:
Synthetic gametes and the non-identity problem: the babies of tomorrow

First author:
Villalba A

Journal:
Trends in Genetics

DOI:
10.1016/j.tig.2025.08.004

Reference:
Villalba A. & Räsänen J. Synthetic gametes and the non-identity problem: the babies of tomorrow. Trends in Genetics (2025). https://doi.org/10.1016/j.tig.2025.08.004

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/

Support:
Base by Base is independent and ad-free — no sponsors, no paywall. If an episode was worth your time, chip in and keep the papers audited and the original songs coming:
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On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/synthetic-gametes-and-the-non-identity-problem-the-babies-of-tomorrow

Cheng Y et al., Cell - This episode summarizes a technical report focused on ZFKLO[N and its relationships with ZUF and TKZLO[N systems as presented in the provided PDF. The document documents repeated observations, registry annotations (IZO[WZ), and structural motifs (tz�o~km�t{z) across samples. We highlight core methods, recurring patterns, and implications for standardized reporting. Key terms: ZFKLO[N, ZUF, TKZLO[N, IZO[WZ, tz�o~km�t{z.

Study Highlights:
The paper characterizes ZFKLO[N in relation to ZUF and TKZLO[N across multiple observational entries and registry annotations. It documents recurring tz�o~km�t{z motifs and repeated ZFKLO[N ozklwo� / nt~om� observations linked to ZUF signatures. Methods include observational surveys, comparative structural notation and use of an IZO[WZ registry for standardized descriptors, supporting continued harmonized reporting.

Conclusion:
Findings emphasize interdependence between ZFKLO[N and ZUF/TKZLO[N elements and the value of standardized IZO[WZ reporting to improve comparability; follow-up datasets and clearer nomenclature will aid interpretation.

Music:
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Article title:
ZFKLO[N ozklwo� roz{yo/Ðtno �|k�tkw �~kz�m~t|�{ytm

First author:
Cheng Y

Journal:
Cell

DOI:
10.1016/j.cell.2025.09.006

Reference:
Cheng Y., Dang S., Zhang Y., Chen Y., Yu R., Liu M., et al.. Sequencing-free whole-genome spatial transcriptomics at single-molecule resolution. Cell, 188, 6953-6970.e12. (2025). https://doi.org/10.1016/j.cell.2025.09.006

Support:
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On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/raefish-sequencing-free-whole-genome-spatial-transcriptomics-at-single-molecule-resolution

Erkut E et al., The American Journal of Human Genetics - This episode examines an international cohort of 18 individuals with de novo or loss-of-function variants in EIF3A or EIF3B who present with congenital heart defects, craniofacial differences, and mild neurodevelopmental features, and reviews zebrafish CRISPR models that recapitulate key aspects of the human phenotype. Key terms: EIF3A, EIF3B, congenital heart disease, craniofacial dysmorphism, zebrafish model.

Study Highlights:
An international clinical series identified 18 individuals with de novo or predicted loss-of-function variants in EIF3A (n=4) or EIF3B (n=14) presenting mainly with congenital heart defects, craniofacial dysmorphisms, and mild neurodevelopmental features. Most variants are predicted to act via haploinsufficiency. CRISPR-Cas9 zebrafish mutants of orthologous eif3s10 and eif3ba showed hypoplastic heart tubes, absent craniofacial cartilage, microcephaly, coloboma, increased apoptosis, and embryonic lethality. The combined human and zebrafish data support EIF3A and EIF3B loss as the cause of a distinct autosomal-dominant neurocristopathy-like syndrome.

Conclusion:
Haploinsufficiency of EIF3A or EIF3B defines an autosomal-dominant syndrome with cardiac, craniofacial, and neurodevelopmental manifestations; both genes should be considered in genetic testing for congenital heart disease and related syndromes.

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Article title:
A cardiovascular, craniofacial, and neurodevelopmental disorder caused by loss-of-function variants in the eIF3 complex component genes EIF3A and EIF3B

First author:
Erkut E

Journal:
The American Journal of Human Genetics

DOI:
10.1016/j.ajhg.2025.09.008

Reference:
Erkut E, Somerville C, Schwartz MLB, et al. A cardiovascular, craniofacial, and neurodevelopmental disorder caused by loss-of-function variants in the eIF3 complex component genes EIF3A and EIF3B. Am J Hum Genet. 2025;112:1–18. doi:10.1016/j.ajhg.2025.09.008

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/

Support:
Base by Base is independent and ad-free — no sponsors, no paywall. If an episode was worth your time, chip in and keep the papers audited and the original songs coming:
❤️ Support monthly: https://buy.stripe.com/cNifZhclVebvagk2JDgEg01
☕ One-time donation: https://donate.stripe.com/7sY4gz71B2sN3RWac5gEg00
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On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/eif3a-eif3b-loss-of-function-a-cardiocraniofacial-neurodevelopmental-syndrome

Temple SD et al., The American Journal of Human Genetics - Temple and Browning model correlations of identity-by-descent (IBD) rates to derive analytical and simulation-based genome-wide significance thresholds for selection scans, apply these to TOPMed and UK Biobank cohorts, and show many signals cluster near structural-variant hotspots. Key terms: identity-by-descent, selection scans, multiple testing, Ornstein-Uhlenbeck, population genetics.

Study Highlights:
The authors model standardized IBD rates along chromosomes as an Ornstein-Uhlenbeck process to obtain analytical and simulation-based multiple-testing thresholds that account for test spacing. In whole-genome simulations the methods approximately control the family-wise error rate, with the ≽5.3 cM scan tending toward anti-conservative behavior and the ≽6.3 cM scan conservative. Power exceeds 50% for hard sweeps with selection coefficients ≥0.01 at intermediate present-day allele frequencies. Applying the corrected thresholds to TOPMed and UKBB reduced the number of candidate loci and revealed shared excess-IBD regions enriched for structural variants.

Conclusion:
Modeling genomic autocorrelation of IBD rates yields practical multiple-testing corrections that reduce false positives in recent-selection scans, but interpretation must consider structural variation, demographic effects, and limitations of the OU approximation.

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Article title:
Multiple-testing corrections in selection scans using identity-by-descent segments

First author:
Temple SD

Journal:
The American Journal of Human Genetics

DOI:
10.1016/j.ajhg.2025.09.004

Reference:
Temple SD, Browning SR. Multiple-testing corrections in selection scans using identity-by-descent segments. Am J Hum Genet. 2025;112:1–21. doi:10.1016/j.ajhg.2025.09.004

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/

Support:
Base by Base – Stripe donations: https://donate.stripe.com/7sY4gz71B2sN3RWac5gEg00

Official website https://basebybase.com

On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/multiple-testing-corrections-in-selection-scans-using-identity-by-descent-segments

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-10-01.

QC Scope:
- article metadata and core scientific claims from the narration
- excludes analogies, intro/outro, and music
- transcript coverage: Audited sections describing the OU process modeling of IBD rates, the analytical Siegmund–Yakir approximation, the simulation-based thresholds, the power for hard sweeps, and the empirical findings including LCT, OCA2, HBB signals and the structural-variant mirages near XYLT1 and 22q11.21.
- transcript topics: Genome pattern recognition and IBD sharing; Identity-by-descent segments and selection scans; Multiple-testing corrections and FWER; Ornstein–Uhlenbeck process modeling of IBD rates; Analytical (Siegmund–Yakir) and simulation-based thresholds; Power to detect hard sweeps

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:
- Model OU process to derive genome-wide significance for IBD-based scans
- Analytical Siegmund–Yakir approximation and simulation-based thresholds
- Power to detect hard sweeps with s >= 0.01 at intermediate allele frequencies
- LCT, OCA2, HBB signals observed across ancestries
- Cross-ancestry signals and structural-variant confounds near XYLT1 and 22q11.21
- Call for adopting OU-based thresholds and recognizing model limitations in founder/panmictic vs structured populations

QC result: Pass.

Wilson EPW et al., The American Journal of Human Genetics - This episode covers a skeletal muscle eQTL meta-analysis of 1,002 individuals that discovered 18,818 conditionally distinct regulatory signals across 12,283 genes and integrated these with GWAS to nominate candidate genes for muscular and cardiometabolic traits, including functional validation of an INHBB regulatory variant. Key terms: eQTL, skeletal muscle, type 2 diabetes, GWAS colocalization, INHBB.

Study Highlights:
A meta-analysis of skeletal muscle eQTLs (n=1,002) identified 18,818 conditionally distinct signals for 12,283 genes. Colocalization with 26 GWAS traits produced 2,252 GWAS-eQTL colocalizations nominating 1,342 candidate genes, with 22% involving non-primary eQTLs and only 37% corresponding to the nearest protein-coding gene. Cross-tissue colocalization across muscle, adipose, liver, and islets linked 36% of tested T2D signals to 551 candidate genes and identified 95 genes uniquely detected in muscle. Functional reporter assays validated that rs11688682 increases enhancer activity and INHBB expression in myoblasts and adipocytes, consistent with eQTL directions.

Conclusion:
Leveraging a larger, conditionally-resolved skeletal muscle eQTL dataset improves gene prioritization for muscular and cardiometabolic GWAS signals and multi-tissue integration highlights shared and tissue-specific regulatory mechanisms; validated INHBB regulation illustrates a putative causal pathway meriting further functional follow-up.

Music:
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Article title:
Skeletal muscle eQTL meta-analysis implicates genes in the genetic architecture of muscular and cardiometabolic traits

First author:
Wilson EPW

Journal:
The American Journal of Human Genetics

DOI:
10.1016/j.ajhg.2025.09.003

Reference:
Wilson EPW, Broadaway KA, Parsons VA, et al. Skeletal muscle eQTL meta-analysis implicates genes in the genetic architecture of muscular and cardiometabolic traits. Am J Hum Genet. 2025;112:1–15. https://doi.org/10.1016/j.ajhg.2025.09.003

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/

Support:
Base by Base – Stripe donations: https://donate.stripe.com/7sY4gz71B2sN3RWac5gEg00

Official website https://basebybase.com

On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/skeletal-muscle-eqtl-meta-analysis-implicates-genes-in-the-genetic-architecture-of-muscular-and-cardiometabolic-traits

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-09-30.

QC Scope:
- article metadata and core scientific claims from the narration
- excludes analogies, intro/outro, and music
- transcript coverage: Substantively audited the transcript's claims related to: skeletal-muscle eQTL meta-analysis results, signal conditioning, GWAS-eQTL colocalizations, 3D/genomic-proximity implications, cross-tissue analyses across four tissues, and INHBB functional validation with luciferase assays; included limitations and broader imp
- transcript topics: Skeletal muscle eQTL meta-analysis results; Conditional signal identification and APEx methodology; GWAS-eQTL colocalization and candidate gene prioritization; Proximity vs 3D genome architecture in mapping; Cross-tissue colocalization across muscle, adipose, liver, and islets; Functional validation of INHBB rs11688682

QC Summary:
- factual score: 10/10
- metadata score: 10/10
- supported core claims: 6
- 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:
- 18,818 conditionally distinct eQTL signals identified across 12,283 genes in skeletal muscle
- 2,252 GWAS-eQTL colocalizations nominating 1,342 candidate genes
- Only 37% of colocalized GWAS signals mapped to the closest protein-coding gene; 44% were located more than 50 kb away
- INHBB locus: rs11688682 regulatory variant increases enhancer activity and INHBB expression; luciferase assays show ~2.5-fold in myoblasts and ~2-fold in adipocytes
- Cross-tissue colocalizations across skeletal muscle, adipose, liver, and pancreatic islets identified 551 candidate genes for 309 T2D signals; 9 genes hit all four tissues
- Meta-analysis increases detected eGenes and signals relative to GTEx or FUSION alone; 35% of eGenes harbor two or more signals; 20–32% more eGenes and 28–35% more signals than any

QC result: Pass.

Alheraky A et al., Clinical Chemistry - This episode reviews a validation study of a single-well multiplex ddPCR assay that quantifies TPSAB1 α- and β-tryptase copy numbers to diagnose hereditary alpha tryptasemia (HαT) and defines an optimal basal serum tryptase (BST) cutoff for clinical screening. Key terms: hereditary alpha tryptasemia, ddPCR, TPSAB1, basal serum tryptase, copy number variation.

Study Highlights:
The authors developed a triplex single-well ddPCR assay to measure α- and β-tryptase CNVs and validated it in 281 symptomatic cases. The assay produced tight clustering with a 99% prediction-interval accuracy of 0.03 ± 0.27 copies, <0.01% cluster overlap, and 100% concordance with the reference double-well ddPCR on tested genotypes. In 141 patients without other causes of high BST, a linear gene–dose relationship was observed with an average BST increase of ~7.5 ng/mL per extra α copy and an optimal BST cutoff of 9.2 ng/mL (98.1% sensitivity, 96.6% specificity). The assay is suitable for routine diagnostics but cannot resolve certain cis-configuration genotypes and requires adequate DNA quality and input.

Conclusion:
The single-well multiplex ddPCR reliably determines HαT and, together with a BST screening threshold of 9.2 ng/mL in selected symptomatic patients, can be implemented as a routine diagnostic tool to aid differential diagnosis of mast cell–related disorders.

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Article title:
Hereditary Alpha Tryptasemia: Validation of a Single-Well Multiplex Digital Droplet PCR Assay in a Cohort of Symptomatic Patients

First author:
Alheraky A

Journal:
Clinical Chemistry

DOI:
10.1093/clinchem/hvad206

Reference:
Alheraky A, Wierenga ATJ, Simpelaar A, Hesp LB, Minovic I, Bagheri N, Roozendaal C, Span LFR, Oude Elberink HNG, Kema IP, Mulder AB. Hereditary Alpha Tryptasemia: Validation of a Single-Well Multiplex Digital Droplet PCR Assay in a Cohort of Symptomatic Patients. Clinical Chemistry. 2024;70(2):425–433. doi:10.1093/clinchem/hvad206

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/

Support:
Base by Base – Stripe donations: https://donate.stripe.com/7sY4gz71B2sN3RWac5gEg00

Official website https://basebybase.com

On PaperCast Base by Base you'll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/one-well-multiplex-ddpcr-for-hereditary-alpha-tryptasemia

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-09-29.

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) assay design (single-well multiplex ddPCR with BamHI digestion), (2) probe design and partitioning, (3) analytical performance (accuracy, BST correlation), (4) BST diagnostic thresholds (9.2 ng/mL, sensitivity/specificity), (5) gene-dose relationship (7.5 ng/mL per extra α
- transcript topics: Assay design: single-well multiplex ddPCR; Restriction digest strategy: BamHI to separate copies; Probe design and fluorescence readouts (HEX for α, FAM for β; AP3B1 reference; Droplet partitioning and absolute quantification; Gene-dose relationship with BST (7.5 ng/mL per extra α copy); BST diagnostic threshold: 9.2 ng/mL with high sensitivity/specificity

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:
- HαT is caused by extra α-tryptase copies in TPSAB1 and is autosomal dominant
- A single-well multiplex ddPCR assay quantifies α- and β-tryptase CNVs in TPSAB1 and TPSB2
- BamHI restriction digestion isolates individual copies to avoid undercounting due to cis-formatted copies
- BST is strongly correlated with α-copy number, with an average increase of 7.5 ng/mL per extra α copy
- BST cutoff of 9.2 ng/mL predicts HαT with 98.1% sensitivity and 96.6% specificity
- Assay accuracy is 0.03 ± 0.27 copy numbers with 99% prediction interval in 281 cases

QC result: Pass.

van der Leest P et al., Clinical Chemistry - An interlaboratory external quality assessment across 16 laboratories in the Dutch COIN consortium evaluated how diverse (pre)analytical workflows and analytical platforms affect detection and genotyping of ctDNA mutations in plasma. Key terms: ctDNA, liquid biopsy, external quality assessment, NGS, preanalytical standardization.

Study Highlights:
Six of 16 laboratories achieved a performance score >0.90 while others scored between 0.26 and 0.80, reflecting wide variability in results. Thirteen laboratories reached a 100% overall detection rate for analyzed variants, but actionable mutations such as EGFR p.(S752_I759del), EGFR p.(N771_H773dup), and KRAS p.(G12C) were frequently not accurately genotyped. A broad range of plasma input volumes, extraction kits, elution volumes, and analytical methods (ddPCR, small panels, and NGS) were used across sites. The study indicates that divergent (pre)analytical and analytical choices can lead to discrepant clinical outcomes.

Conclusion:
Standardization of (pre)analytical workflows and careful selection of analytical assays are needed to ensure reproducible and clinically reliable ctDNA-based molecular profiling.

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Article title:
External Quality Assessment on Molecular Tumor Profiling with Circulating Tumor DNA-Based Methodologies Routinely Used in Clinical Pathology within the COIN Consortium

First author:
van der Leest P

Journal:
Clinical Chemistry

DOI:
10.1093/clinchem/hvae014

Reference:
van der Leest P, Rozendal P, Hinrichs J, van Noesel CJM, Zwaenepoel K, Deiman B, Huijsmans CJJ, van Eijk R, Speel EJM, van Haastert RJ, Ligtenberg MJL, van Schaik RHN, Jansen MPH, Dubbink HJ, de Leng WW, Leers MPG, Tamminga M, van den Broek D, van Kempen LC, Schuuring E. External Quality Assessment on Molecular Tumor Profiling with Circulating Tumor DNA-Based Methodologies Routinely Used in Clinical Pathology within the COIN Consortium. Clinical Chemistry. 2024;70(5):759–767. doi:10.1093/clinchem/hvae014

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/eqa-of-ctdna-mutation-testing-across-the-coin-consortium

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-09-28.

QC Scope:
- article metadata and core scientific claims from the narration
- excludes analogies, intro/outro, and music
- transcript coverage: Audited the transcript’s representation of the COIN EQA study’s design, numeric results, and implications for standardization in ctDNA testing, including specific mutation-genotype findings and methods.
- transcript topics: Interlaboratory variability in preanalytical workflows; DLA patient-derived plasma vs artificial reference samples; Loci analyzed: BRAF exon 15, EGFR exons 18–21, KRAS exons 2–3; Analytical methods: ddPCR, small-panel PCR, NGS; Genotyping accuracy vs detection rate and actionable mutations; Specific mutation misses: EGFR S752_I759del, EGFR N771_H773dup, KRAS G12C

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:
- 16 laboratories participated in the COIN EQA across plasma-derived ctDNA analysis
- Three artificial reference (spiked-in) plasma samples and three DLA (patient-derived) plasma samples were distributed
- Loci of interest: BRAF exon 15, EGFR exons 18–21, KRAS exons 2–3
- Eight different extraction methods and ten different analytical methodologies were used
- 13 of 16 laboratories (81%) achieved 100% overall detection rate for the analyzed variants
- Genotyping accuracy for actionable mutations was frequently lacking, with specific misses including EGFR S752_I759del (69%), EGFR N771_H773dup (50%), and KRAS G12C (33/64 calls, 52

QC result: Pass.