BACKGROUND: Inherited Long QT Syndrome (LQTS) is a cardiac channelopathy associated with a high risk of sudden death. The prevalence has been estimated at close to 1:2000. Due to large cohorts to investigate, the size of the 3 prevalent mutated genes, and the presence of a large spectrum of private mutations, mutational screening requires an extremely sensitive and specific scanning method. METHODS: Efficiency of high resolution melting (HRM) analysis was evaluated for the most prevalent LQTS-causing genes (KCNQ1, KCNH2) using control DNAs and DNAs carrying previously identified gene variants. A cohort of 34 patients with a suspicion of LQTS was further blindly screened. To evaluate HRM sensitivity, this cohort was also screened using an optimized DHPLC strategy. RESULTS: HRM analysis was successfully optimized for KCNQ1 but optimisation of KCNH2 was more laborious as only 3 KCNH2 exons could be finally optimized. Remaining KCNH2 exons were analysed by direct sequencing. This molecular approach, which combined HRM and direct sequencing, was applied on the cohort of 34 cases and 9 putative mutations were identified. Using this approach, molecular investigation was completed faster and cheaper than using DHPLC strategy. CONCLUSIONS: This HRM/sequencing procedure represents an inexpensive, highly sensitive and high-throughput method to allow identification of mutations in the coding sequences of prevalent LQTS genes.
BACKGROUND:Inherited Long QT Syndrome (LQTS) is a cardiac channelopathy associated with a high risk of sudden death. The prevalence has been estimated at close to 1:2000. Due to large cohorts to investigate, the size of the 3 prevalent mutated genes, and the presence of a large spectrum of private mutations, mutational screening requires an extremely sensitive and specific scanning method. METHODS: Efficiency of high resolution melting (HRM) analysis was evaluated for the most prevalent LQTS-causing genes (KCNQ1, KCNH2) using control DNAs and DNAs carrying previously identified gene variants. A cohort of 34 patients with a suspicion of LQTS was further blindly screened. To evaluate HRM sensitivity, this cohort was also screened using an optimized DHPLC strategy. RESULTS: HRM analysis was successfully optimized for KCNQ1 but optimisation of KCNH2 was more laborious as only 3 KCNH2 exons could be finally optimized. Remaining KCNH2 exons were analysed by direct sequencing. This molecular approach, which combined HRM and direct sequencing, was applied on the cohort of 34 cases and 9 putative mutations were identified. Using this approach, molecular investigation was completed faster and cheaper than using DHPLC strategy. CONCLUSIONS: This HRM/sequencing procedure represents an inexpensive, highly sensitive and high-throughput method to allow identification of mutations in the coding sequences of prevalent LQTS genes.