Reflex RNA Sequencing for Variant Classification Improvement

This technical report discusses the integration of reflex RNA sequencing (RNAseq) into the clinical workflow of exome and genome sequencing (ES/GS) to enhance the classification of variants of uncertain significance (VUS). The report outlines the methodology used, including the extraction of DNA and RNA from peripheral blood, and the criteria for variant inclusion based on their association with patient phenotypes and predicted splicing effects. The results indicate that the addition of RNAseq significantly improved the reclassification of VUS to likely pathogenic variants, with a notable 50% upgrade rate observed in the cases studied. Clinical outcomes are documented, demonstrating how RNAseq provided timely answers that traditional testing methods could not, thereby facilitating better patient management. The findings suggest that RNAseq is a valuable tool in the context of rare diseases, where accurate variant classification is critical for patient treatment and monitoring.