2023 ACMG Annual Clinical Genetics Meeting Digital Edition: Poster Gallery-A Comprehensive PCR/Nanopore Based Panel for CFTR Population Carrier Screening that Resolves Multiple Classes of Clinically-Relevant Variants

A Comprehensive PCR/Nanopore Based Panel for CFTR Population Carrier Screening that Resolves Multiple Classes of Clinically-Relevant Variants
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Pdf Summary
The study aimed to develop a PCR/nanopore assay for carrier screening of cystic fibrosis (CF) that can detect a wide range of pathogenic variants in the CFTR gene. The researchers obtained genomic DNA samples from cell lines and whole blood samples, which were amplified, barcoded, and sequenced using the Oxford Nanopore Technologies (ONT) MinION platform. The performance of ONT basecalling models and variant callers was evaluated for SNV and indel detection, and bespoke algorithms were developed to detect large exon deletions. The assay was compared to other methods such as Sanger sequencing. The results showed that the PCR/nanopore assay accurately detected SNVs, indels, and large exon deletions with 100% accuracy in cell-line samples and more than 93% agreement in whole blood samples compared to orthogonal data. The assay also showed high sensitivity and positive predictive value for pathogenic SNV and indel identification. Additionally, the assay successfully identified and phased variants adjacent to homopolymers and polyT/TG repeats. The study concluded that the PCR/nanopore assay, using the latest ONT kits and software, can effectively detect multiple challenging variants in the CFTR gene for CF carrier screening. The combination of PCR enrichment and nanopore sequencing provides a comprehensive and scalable option for carrier screening, especially as professional guidelines recommend increasing the number of variants considered for diverse populations. The researchers also noted that long-read sequencing with nanopore technology can overcome the limitations of short-read sequencing in detecting and phasing complex variants. The PCR/nanopore approach has the potential to expand the repertoire of reported variants relevant to CF carrier screening. Overall, the study demonstrates the feasibility of using PCR enrichment and nanopore sequencing for accurate and comprehensive CFTR variant detection in carrier screening. The assay has the potential to improve test equity and address the needs of diverse populations. Further development and validation of the assay are needed before its future availability and performance can be determined.

Asset Subtitle

Presenting Author - Monica P. Roberts, MS;

Meta Tag

Bioinformatics

Genetic Testing

Genome sequencing

Identification of Disease Genes

Respiratory System

Risk Assessment

Sequencing

Variant Detection

Presenting Author Monica P. Roberts, MS

Keywords

PCR/nanopore assay

carrier screening

cystic fibrosis

CFTR gene

pathogenic variants

genomic DNA samples

Oxford Nanopore Technologies

ONT MinION platform

SNV detection

indel detection