title: Enhanced diagnostic yield in Meckel-Gruber and Joubert syndrome through exome sequencing supplemented with split-read mapping.
abstract: BACKGROUND
  The widespread adoption of high-throughput sequencing technologies by genetic diagnostic laboratories has enabled significant expansion of their testing portfolios. Rare autosomal recessive conditions have been a particular focus of many new services. Here we report a cohort of 26 patients referred for genetic analysis of Joubert (JBTS) and Meckel-Gruber (MKS) syndromes, two clinically and genetically heterogeneous neurodevelopmental conditions that define a phenotypic spectrum, with MKS at the severe end.
  
  
  METHODS
  Exome sequencing was performed for all cases, using Agilent SureSelect v5 reagents and Illumina paired-end sequencing. For two cases medium-coverage (9×) whole genome sequencing was subsequently undertaken.
  
  
  RESULTS
  Using a standard analysis pipeline for the detection of single nucleotide and small insertion or deletion variants, molecular diagnoses were confirmed in 12 cases (4%). Seeking to determine whether our cohort harboured pathogenic copy number variants (CNV), in JBTS- or MKS-associated genes, targeted comparative read-depth analysis was performed using FishingCNV. These analyses identified a putative intragenic AHI1 deletion that included three exons spanning at least 3.4 kb and an intergenic MPP4 to TMEM237 deletion that included exons spanning at least 21.5 kb. Whole genome sequencing enabled confirmation of the deletion-containing alleles and precise characterisation of the mutation breakpoints at nucleotide resolution. These data were validated following development of PCR-based assays that could be subsequently used for "cascade" screening and/or prenatal diagnosis.
  
  
  CONCLUSIONS
  Our investigations expand the AHI1 and TMEM237 mutation spectrum and highlight the importance of performing CNV screening of disease-associated genes. We demonstrate a robust increasingly cost-effective CNV detection workflow that is applicable to all MKS/JBTS referrals.
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. c.m.watson@leeds.ac.uk.|School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. c.m.watson@leeds.ac.uk.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Watson
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. l.a.crinnion@leeds.ac.uk.|School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. l.a.crinnion@leeds.ac.uk.
authors.forename: Laura A
authors.initials: LA
authors.lastname: Crinnion
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. ianberry@nhs.net.
authors.forename: Ian R
authors.initials: IR
authors.lastname: Berry
authors.affiliation: School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. s.harrison@leeds.ac.uk.
authors.forename: Sally M
authors.initials: SM
authors.lastname: Harrison
authors.affiliation: School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. c.lascelles@leeds.ac.uk.
authors.forename: Carolina
authors.initials: C
authors.lastname: Lascelles
authors.affiliation: School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. umaan@leeds.ac.uk.
authors.forename: Agne
authors.initials: A
authors.lastname: Antanaviciute
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. ruth.charlton1@nhs.net.
authors.forename: Ruth S
authors.initials: RS
authors.lastname: Charlton
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. angus.dobbie@nhs.net.
authors.forename: Angus
authors.initials: A
authors.lastname: Dobbie
authors.affiliation: School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. i.m.carr@leeds.ac.uk.
authors.forename: Ian M
authors.initials: IM
authors.lastname: Carr
authors.affiliation: Yorkshire Regional Genetics Service, St. James's University Hospital, Leeds, LS9 7TF, UK. d.t.bonthron@leeds.ac.uk.|School of Medicine, University of Leeds, St. James's University Hospital, Leeds, LS9 7TF, UK. d.t.bonthron@leeds.ac.uk.
authors.forename: David T
authors.initials: DT
authors.lastname: Bonthron
mesh_terms: D000015:Abnormalities, Multiple / Q000175:diagnosis / Q000235:genetics; D000483:Alleles; D002531:Cerebellum / Q000002:abnormalities*; D002874:Chromosome Mapping; D002925:Ciliary Motility Disorders / Q000175:diagnosis* / Q000235:genetics*; D015331:Cohort Studies; D056915:DNA Copy Number Variations; D004677:Encephalocele / Q000175:diagnosis* / Q000235:genetics*; D059472:Exome; D005091:Exons; D005124:Eye Abnormalities / Q000175:diagnosis / Q000235:genetics; D005820:Genetic Testing; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D052177:Kidney Diseases, Cystic / Q000175:diagnosis / Q000235:genetics; D007690:Polycystic Kidney Diseases / Q000175:diagnosis* / Q000235:genetics*; D011296:Prenatal Diagnosis; D012160:Retina / Q000002:abnormalities*; D012174:Retinitis Pigmentosa; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion