1. Design and application of a 23-gene panel by next-generation sequencing for inherited coagulation bleeding disorders
- Author
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Maria Luisa Lozano, Luis Javier García-Frade, Ramón García-Sanz, M. del Rey, Rosa Fisac, M. J. Cebeira, Rafael Ramos, José María Bastida, José Ramón González-Porras, C. Aguilera, M. E. Sarasquete, J.M. Hernández-Rivas, Marcos González-Díaz, Emilia Pardal, M. E. Fontecha, B. Pérez, Susana Riesco, Rocío Benito, J. M. Martin-Antorán, Nuria Bermejo, M. C. Mendoza, Carlos Aguilar, María Paz Martínez, M. R. Cardesa, and L. A. Silvestre
- Subjects
Adult ,Male ,0301 basic medicine ,Haemophilia ,Adolescent ,Genotype ,Genetic counseling ,Mutation, Missense ,Prenatal diagnosis ,030204 cardiovascular system & hematology ,Biology ,DNA sequencing ,Inherited rare bleeding disorders ,Frameshift mutation ,Fibrinogen disorders ,Young Adult ,03 medical and health sciences ,symbols.namesake ,Blood Coagulation Disorders, Inherited ,0302 clinical medicine ,medicine ,Humans ,Genetic Testing ,Child ,Frameshift Mutation ,Genotyping ,Genetic Association Studies ,Genetics (clinical) ,Genetic testing ,Genetics ,Sanger sequencing ,medicine.diagnostic_test ,High-Throughput Nucleotide Sequencing ,Infant ,DNA ,Sequence Analysis, DNA ,Hematology ,General Medicine ,Middle Aged ,Inherited coagulation factor deficiencies ,030104 developmental biology ,Child, Preschool ,Next-generation sequencing ,symbols ,Female ,Molecular diagnosis ,Gene Deletion - Abstract
[Introduction]: Molecular testing of Inherited bleeding coagulation disorders (IBCDs) not only offers confirmation of diagnosis but also aids in genetic counselling, prenatal diagnosis and in certain cases genotype–phenotype correlations are important for predicting the clinical course of the disease and to allow tailor-made follow-up of individuals. Until recently, genotyping has been mainly performed by Sanger sequencing, a technique known to be time consuming and expensive. Currently, next-generation sequencing (NGS) offers a new potential approach that enables the simultaneous investigation of multiple genes at manageable cost. [Aim]: The aim of this study was to design and to analyse the applicability of a 23-gene NGS panel in the molecular diagnosis of patients with IBCDs. [Methods]: A custom target enrichment library was designed to capture 31 genes known to be associated with IBCDs. Probes were generated for 296 targets to cover 86.3 kb regions (all exons and flanking regions) of these genes. Twenty patients with an IBCDs phenotype were studied using NGS technology. [Results]: In all patients, our NGS approach detected causative mutations. Twenty-one pathogenic variants were found; while most of them were missense (18), three deletions were also identified. Six novel mutations affecting F8, FGA, F11, F10 and VWF genes, and 15 previously reported variants were detected. NGS and Sanger sequencing were 100% concordant. Conclusion: Our results demonstrate that this approach could be an accurate, reproducible and reliable tool in the rapid genetic diagnosis of IBCDs.
- Published
- 2016
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