Integrative approach for delineating structural variants using optical genome mapping and long-read genome sequencing

Structural variants (SVs) are genomic rearrangements of 50 bp or more and contribute substantially to human genetic disorders. They are often incompletely resolved using cytogenetic and short-read sequencing approaches. Emerging technologies such as long-read sequencing (LRS) and optical genome mapping (OGM) provide complementary advantages for SV detection. This study aimed to evaluate the combined utility of OGM and LRS for the precise delineation of clinically relevant chromosomal rearrangements.

Five patients with suspected chromosomal abnormalities were investigated using karyotyping, fluorescence in situ hybridisation (FISH), chromosomal microarray (CMA), and triplet repeat-primed PCR (TP-PCR), followed by integrative analysis using OGM and LRS.

In Case 1 (short stature and recurrent miscarriages), OGM identified two insertions in addition to a balanced translocation, whereas LRS revealed disruptions in the NBAS and VTI1A genes. In Case 2 (Fragile X syndrome), LRS precisely determined the exact size of FMR1 CGG repeats, supported by OGM. In Case 3, LRS resolved inversion breakpoints with associated microdeletions and insertions associated with recurrent miscarriages. In Case 4, OGM identified an additional intrachromosomal rearrangement on chromosome 18. In Case 5, OGM detected an intrachromosomal fusion underlying intellectual disability. Overall, OGM provided genome-wide structural insights, while LRS enabled nucleotide-level breakpoint resolution. Except for case 2, all rearrangements are novel.

Our study concludes that the combined application of technologies enables comprehensive characterisation of complex SVs that remain unresolved by standard methods. This integrative approach improves diagnostic yield, facilitates genotype-phenotype correlation, and supports the implementation in SV detection for patients.