Introduction Clinical Exome Sequencing (ES) serves as a powerful diagnostic tool, enabling the simultaneous detection of single nucleotide variants (SNVs) and copy number variations (CNVs) in a single workflow. However, accurate CNV calling from exome data remains challenging due to factors such as the reliance on a panel of normals, uneven coverage in certain regions, and the presence of pseudogenes. To address these limitations, backbone-probe-enhanced exome panels have been developed. By incorporating enhanced target capture and a robust analytical pipeline, this approach offers improved CNV detection and presents a compelling alternative to conventional techniques like chromosomal microarrays (CMA). Methods Twist Bioscience’s Twist Exome 2.0 Plus Comprehensive Exome capture panel incorporates 25kb, 50kb, and 100kb interval backbone probes evenly distributed across the genome to enhance coverage. Simultaneously, Geneyx has adopted Illumina’s DRAGEN CNV caller, optimized for target-capture next-generation sequencing (NGS) data, to detect CNV events ranging from single-exon variations to whole-chromosome anomalies. This study evaluated the combined performance of the Twist Exome 2.0 Plus panel and the Geneyx-implemented DRAGEN pipeline in detecting CNVs within the Coriell CNVPANEL01 reference set, consisting of 43 “genome in a bottle” samples. Results The integration of the enhanced capture panel with the DRAGEN CNV pipeline achieved ∼100% concordance in detecting known CNV events. This included both large-scale events (>10kb), comparable to those identified by CMA, and smaller single-exon events (