From sample to clinical insight: a review of exome sequencing in disease diagnostics

Exome sequencing (ES) has transformed genomic research and clinical diagnostics by enabling precise identification of disease-associated variants within protein-coding regions, which, while representing a minority of the genome, include many well-characterized pathogenic mutations. This review provides a comprehensive overview of ES methodology, data analysis pipelines, clinical relevance, and ethical considerations. We describe the ES workflow from DNA extraction and library preparation to target enrichment, sequencing to ES data analysis. We have also evaluated major capture technologies and sequencing platforms, including short-read and emerging long-read systems. Furthermore, we discuss computational analysis tools such as GATK, FreeBayes, DeepVariant, and Platypus, and strategies to improve accuracy through rigorous quality control, coverage optimization, and orthogonal validation. Beyond rare disease and cancer genomics, ES has expanded into pharmacogenomics, population-scale studies, and integrative multi-omics frameworks that combine transcriptomic and proteomic data to enhance functional interpretation. We highlight actionable examples such as CYP2C19 variants influencing clopidogrel metabolism, illustrating ES's growing role in personalized medicine. Challenges (including variant interpretation complexity, false positives, and data standardization) are critically discussed. The review also addresses ethical, legal, and social dimensions of ES, including informed consent, data privacy, incidental findings, and adherence to ACMG, HIPAA, and GDPR. Finally, we outline future directions emphasizing machine learning-based variant prioritization, single-cell sequencing integration, and scalable bioinformatics infrastructures to enhance accuracy and clinical translation. Collectively, these developments position ES as a pivotal tool bridging genomic discovery, disease diagnostics, and precision healthcare in the era of personalized medicine.