title: Targeted next generation sequencing for molecular diagnosis of Usher syndrome.
abstract: BACKGROUND
  Usher syndrome is an autosomal recessive disease that associates sensorineural hearing loss, retinitis pigmentosa and, in some cases, vestibular dysfunction. It is clinically and genetically heterogeneous. To date, 10 genes have been associated with the disease, making its molecular diagnosis based on Sanger sequencing, expensive and time-consuming. Consequently, the aim of the present study was to develop a molecular diagnostics method for Usher syndrome, based on targeted next generation sequencing.
  
  
  METHODS
  A custom HaloPlex panel for Illumina platforms was designed to capture all exons of the 10 known causative Usher syndrome genes (MYO7A, USH1C, CDH23, PCDH15, USH1G, CIB2, USH2A, GPR98, DFNB31 and CLRN1), the two Usher syndrome-related genes (HARS and PDZD7) and the two candidate genes VEZT and MYO15A. A cohort of 44 patients suffering from Usher syndrome was selected for this study. This cohort was divided into two groups: a test group of 11 patients with known mutations and another group of 33 patients with unknown mutations.
  
  
  RESULTS
  Forty USH patients were successfully sequenced, 8 USH patients from the test group and 32 patients from the group composed of USH patients without genetic diagnosis. We were able to detect biallelic mutations in one USH gene in 22 out of 32 USH patients (68.75%) and to identify 79.7% of the expected mutated alleles. Fifty-three different mutations were detected. These mutations included 21 missense, 8 nonsense, 9 frameshifts, 9 intronic mutations and 6 large rearrangements.
  
  
  CONCLUSIONS
  Targeted next generation sequencing allowed us to detect both point mutations and large rearrangements in a single experiment, minimizing the economic cost of the study, increasing the detection ratio of the genetic cause of the disease and improving the genetic diagnosis of Usher syndrome patients.
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. majoaparisi@gmail.com.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. majoaparisi@gmail.com.
authors.forename: María J
authors.initials: MJ
authors.lastname: Aparisi
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. elenaller@yahoo.es.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. elenaller@yahoo.es.
authors.forename: Elena
authors.initials: E
authors.lastname: Aller
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. c.fustergarcia@gmail.com.
authors.forename: Carla
authors.initials: C
authors.lastname: Fuster-García
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. gema.garcia@inserm.fr.|CHU Montpellier, Laboratoire de Génétique Moléculaire and Inserm, U827, Montpellier, F-34000, France. gema.garcia@inserm.fr.
authors.forename: Gema
authors.initials: G
authors.lastname: García-García
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. regina.rodrigo@yahoo.es.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. regina.rodrigo@yahoo.es.
authors.forename: Regina
authors.initials: R
authors.lastname: Rodrigo
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. rafael_vazquez@iislafe.es.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. rafael_vazquez@iislafe.es.
authors.forename: Rafael P
authors.initials: RP
authors.lastname: Vázquez-Manrique
authors.affiliation: CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. FBlancoK@fjd.es.|Servicio de Genética, IIS - Fundación Jiménez Díaz, University Hospital, UAM, Madrid, Spain. FBlancoK@fjd.es.
authors.forename: Fiona
authors.initials: F
authors.lastname: Blanco-Kelly
authors.affiliation: CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. cayuso@fjd.es.|Servicio de Genética, IIS - Fundación Jiménez Díaz, University Hospital, UAM, Madrid, Spain. cayuso@fjd.es.
authors.forename: Carmen
authors.initials: C
authors.lastname: Ayuso
authors.affiliation: CHU Montpellier, Laboratoire de Génétique Moléculaire and Inserm, U827, Montpellier, F-34000, France. anne-francoise.roux@inserm.fr.
authors.forename: Anne-Françoise
authors.initials: AF
authors.lastname: Roux
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. tjaijo@gmail.com.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. tjaijo@gmail.com.
authors.forename: Teresa
authors.initials: T
authors.lastname: Jaijo
authors.affiliation: Grupo de Investigación en Enfermedades Neurosensoriales. Instituto de Investigación Sanitaria IIS-La Fe, Semisótano Escuela de Enfermería, Hospital Universitario La Fe, Avda. Campanar, 21, 46009, Valencia, Spain. millan_jos@gva.es.|CIBER de Enfermedades Raras (CIBERER), Valencia, Spain. millan_jos@gva.es.|Unidad de Genética y Diagnóstico Prenatal, Hospital Universitario y Politécnico La Fe, Valencia, Spain. millan_jos@gva.es.
authors.forename: José M
authors.initials: JM
authors.lastname: Millán
mesh_terms: D005260:Female; D020869:Gene Expression Profiling; D015321:Gene Rearrangement; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008297:Male; D025202:Molecular Diagnostic Techniques; D009154:Mutation; D017354:Point Mutation; D052245:Usher Syndromes / Q000175:diagnosis* / Q000235:genetics