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Over the past years, next-generation sequencing (NGS) technologies revolutionized the possibilities in a broad range of application areas. Also in the field of forensic genetics, NGS continuously gained in importance and attentiveness. A significant number of sudden cardiac deaths (SCD) in the young is due to heritable arrhythmia syndromes emphasizing the need of examining the genetic basis in these cases also with regard to the identification of relatives and/or patients being at risk. As a result, high-throughput methods became of increasing value in molecular autopsy investigations enabling the analysis of a broad spectrum of genes.
Most standard protocols are optimized for high-quality samples and frequently not directly applicable to challenging forensic sample material. In the present study, we intended to examine a comprehensive gene panel associated with SCD and inherited arrhythmogenic disorders. We compared three different hybridization-based library preparation technologies in order to implement a suitable NGS workflow for heterogeneous, forensic as well as diagnostic sample material.
The results obtained indicated, that the Illumina technologies Nextera DNA Flex and TruSeq were compatible with samples exhibiting varying levels of degradation. In comparison, the TruSight method also resulted in good sequencing data, but seemed to be more dependent on DNA integrity. The preparation protocols evaluated in our study are not restricted to molecular autopsy investigations and might be helpful for and transferrable to further forensic research applications.
Background: Alterations in the SCN5A gene encoding the cardiac sodium channel Nav1.5 have been linked to a number of arrhythmia syndromes and diseases including long-QT syndrome (LQTS), Brugada syndrome (BrS) and dilative cardiomyopathy (DCM), which may predispose to fatal arrhythmias and sudden death. We identified the heterozygous variant c.316A > G, p.(Ser106Gly) in a 35-year-old patient with survived cardiac arrest. In the present study, we aimed to investigate the functional impact of the variant to clarify the medical relevance.
Methods: Mutant as well as wild type GFP tagged Nav1.5 channels were expressed in HEK293 cells. We performed functional characterization experiments using patch-clamp technique.
Results: Electrophysiological measurements indicated, that the detected missense variant alters Nav1.5 channel functionality leading to a gain-of-function effect. Cells expressing S106G channels show an increase in Nav1.5 current over the entire voltage window.
Conclusion: The results support the assumption that the detected sequence aberration alters Nav1.5 channel function and may predispose to cardiac arrhythmias and sudden cardiac death.