title: DNA sequence capture and next-generation sequencing for the molecular diagnosis of genetic cardiomyopathies.
abstract: Hypertrophic cardiomyopathy is a relatively frequent disease with a prevalence of 0.2% worldwide and a remarkable genetic heterogeneity, with more than 30 causative genes reported to date. Current PCR-based strategies are inadequate for genomic investigations involving many candidate genes. Here, we report a next-generation sequencing procedure associated with DNA sequence capture that is able to sequence 202 cardiomyopathy-related genes simultaneously. We developed a complementary data analysis pipeline to select and prioritize genetic variants. The overall procedure can screen a large number of target genes simultaneously, thereby potentially revealing new disease-causing and modifier genes. By using this procedure, we analyzed hypertrophic cardiomyopathy patients in a shorter time and at a lower cost than with current procedures. The specificity of the next-generation sequencing-based procedure is at least as good as other techniques routinely used for mutation searching, and the sensitivity is much better. Analysis of the results showed some novel variants potentially involved in the pathogenesis of hypertrophic cardiomyopathy: a missense mutation in MYH7 and a nonsense variant in INS-IGF2 (patient 1), a splicing variant in MYBPC3 and an indel/frameshift variant in KCNQ1 (patient 2), and two concomitant variations in CACNA1C (patient 3). Sequencing of DNA from the three patients within a pool allowed detection of most variants identified in each individual patient, indicating that this approach is a feasible and cost-effective procedure.
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: D'Argenio
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Frisso
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.
authors.forename: Vincenza
authors.initials: V
authors.lastname: Precone
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy.
authors.forename: Angelo
authors.initials: A
authors.lastname: Boccia
authors.affiliation: Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.
authors.forename: Antonella
authors.initials: A
authors.lastname: Fienga
authors.affiliation: Cardiomyopathy and Inherited Heart Disease Clinic, UOC Cardiology, Second University of Naples, Naples, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Pacileo
authors.affiliation: Cardiomyopathy and Inherited Heart Disease Clinic, UOC Cardiology, Second University of Naples, Naples, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Limongelli
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Paolella
authors.affiliation: Cardiomyopathy and Inherited Heart Disease Clinic, UOC Cardiology, Second University of Naples, Naples, Italy.
authors.forename: Raffaele
authors.initials: R
authors.lastname: Calabrò
authors.affiliation: CEINGE-Biotecnologie Avanzate, Naples, Italy; IRCCS-Fondazione SDN, Naples, Italy. Electronic address: salvator@unina.it.
authors.forename: Francesco
authors.initials: F
authors.lastname: Salvatore
mesh_terms: D000293:Adolescent; D001483:Base Sequence; D020746:Calcium Channels, L-Type / Q000235:genetics; D024722:Cardiac Myosins / Q000235:genetics; D002312:Cardiomyopathy, Hypertrophic / Q000175:diagnosis* / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics; D002648:Child; D005260:Female; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D051657:KCNQ1 Potassium Channel / Q000235:genetics; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques; D008969:Molecular Sequence Data; D050506:Mutant Chimeric Proteins / Q000235:genetics; D009154:Mutation; D018995:Myosin Heavy Chains / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D000099163:Myosin Binding Protein C
chemical_list: D020746:Calcium Channels, L-Type; D024722:Cardiac Myosins; D002352:Carrier Proteins; D051657:KCNQ1 Potassium Channel; D050506:Mutant Chimeric Proteins; D018995:Myosin Heavy Chains; D000099163:Myosin Binding Protein C; C079146:CACNA1C protein, human; C584675:INS-IGF2 protein, human; C496909:KCNQ1 protein, human; C501641:MYH7 protein, human