Deciphering the Structural Variants by Long-Read Genome Sequencing: Technology, Applications, and Case Illustrations.

Background: Structural variants (SVs) are defined as genomic variants affecting more than 50 base pairs. They include deletions, insertions, inversions, translocations, tandem repeats, and copy number variations. These SVs contribute significantly to genetic complexity and are involved in human evolution, genetic disorders, and cancer. Over 50% of the SVs cannot be detected due to limitations in methods and technologies. The short-read sequencing technologies (SRSs) are limited in detecting single-nucleotide variants and have limited usage for analysis of complex genomic loci, repeat regions, and phasing.

Summary: The advent of long-read sequencing (LRS) technologies, such as Oxford Nanopore and PacBio, has revolutionized SV detection. These platforms enable the accurate characterization of diverse variant types, ranging from simple deletions to complex chromothripsis events, and support de novo assembly, haplotype phasing, and the resolution of repetitive or structurally complex genomic regions. One major outcome is the completion of the telomere-to-telomere human reference genome. This review summarizes recent advances in LRS for SV detection, including sequencing platforms, bioinformatic tools, data analysis, and validation strategies. The clinical applications, particularly in the diagnosis of rare diseases, are illustrated with two cases that were successfully resolved using both LRS approaches.

Key message: LRS can overcome the limitations of SRS in SV detection, providing more accurate insights into genome disorders. It enables the detection of repeat and difficult-to-resolve regions of the genome and facilitates clinical diagnoses to base-level breakpoint detection. Despite challenges such as high cost, data interpretation, and clinical linking, continued advancements are elevating LRS as an invaluable tool in precision genomic medicine.