Mutation Detection in Congenital Long QT Syndrome: Cardiac Channel Gene Screen Using PCR, dHPLC, and Direct DNA Sequencing

Within the field of molecular cardiac electrophysiology, the previous decade of research elucidated the fundamental genetic substrate underlying many arrhythmogenic disorders such as long QT syndrome (LQTS), catecholaminergic polymorphic ventricular tachycardia (CPVT), Andersen-Tawil syndrome, Brugada Syndrome, and Timothy syndrome. In addition, the genetic basis for cardiomyopathic processes vulnerable to sudden arrhythmic death-hypertrophic cardiomyopathy, dilated cardiomyopathy, and arrhythmogenic right ventricular cardiomyopathy-are understood now in greater detail. The majority of congenital LQTS is understood as a primary cardiac channelopathy that often but not always provides evidence of its presence via a prolonged QT interval on the 12-lead surface electrocardiogram. To date, more than 300 mutations have been identified in five genes encoding key ion channel sub units involved in the orchestration of the heart's action potential. LQTS genetic testing has been performed in research laboratories over the past decade, relying on the techniques of PCR, an intermediate mutation analysis platform such as single-stranded conformation polymorphism (SSCP) or denaturing high-performance liquid chromatography (dHPLC), and subsequent direct DNA sequencing to elucidate the genetic underpinnings of this disorder. Presently, LQTS genetic testing is a clinically available molecular diagnostic test that provides comprehensive open reading frame/splice site mutational analysis via high-throughput DNA sequencing. This chapter will focus on LQTS genetic testing employing the techniques of genomic DNA isolation from peripheral blood, exon-specific PCR amplification, dHPLC hetero-duplex analysis, and direct DNA sequencing.