Addressing the diagnostic odyssey beyond variant calling: a perspective on long-read sequencing and context-aware genome interpretation

Short-read exome and genome sequencing have transformed not only clinical genetics but also many other medical specialties that rely on genomic information. Yet, in clinical genetics, many individuals remain undiagnosed because the relevant disease mechanism cannot always be represented as an isolated variant at a single genomic coordinate. Phase, structural configuration, repeat composition, paralog assignment, methylation, and transcript consequences can each influence whether a variant is pathogenic. Long-read sequencing preserves molecular continuity across kilobases of native DNA and, in separate, complementary assays, across full-length RNA transcripts. Its clinical value includes both detecting previously missed variation and reconstructing the molecular configuration of detected findings. In this Perspective, we argue that clinical genomics should increasingly treat haplotypes, structural alleles, repeat alleles, and allele-specific regulatory states as primary diagnostic objects. Using a composite unresolved recessive-disease scenario, we place this perspective within the diagnostic odyssey, discuss health economics and outcomes research (HEOR) considerations, propose a context-aware workflow, and define priorities for validation and evidence generation. We identify potential value under reanalysis as a hypothesis requiring economic evaluation and argue that most current clinical databases are not designed to adequately represent the configurations these assays produce. Long-read sequencing need not universally replace short-read testing; its most compelling role is in settings where preserving molecular continuity changes the clinical meaning and downstream value of the result.