title: Targeted Droplet-Digital PCR as a Tool for Novel Deletion Discovery at the DFNB1 Locus.
abstract: Pathogenic variants at the DFNB1 locus encompassing the GJB2 and GJB6 genes account for 50% of autosomal-recessive, congenital nonsyndromic hearing loss in the United States. Most cases are caused by sequence variants within the GJB2 gene, but a significant number of DFNB1 patients carry a large deletion (GJB6-D13S1830) in trans with a GJB2 variant. This deletion lies upstream of GJB2 and was shown to reduce GJB2 expression by disrupting unidentified regulatory elements. First-tier genetic testing for hearing loss includes GJB2 sequence and GJB6-D13S1830 deletion analysis; however, several other deletions in this locus, each with distinct breakpoints, have been reported in DFNB1 patients and are missed by current panels. Here, we report the development of a targeted droplet digital polymerase chain reaction-based assay for comprehensive copy-number analysis at the DFNB1 locus that detects all deletions reported to date. This assay increased detection rates in a multiethnic cohort of 87 hearing loss patients with only one identified pathogenic GJB2 variant. We identify two deletions, one of which is novel, in two patients (2/87 or 2.3%), suggesting that other pathogenic deletions at the DFNB1 locus may be missed. Mapping the assayed DFNB1 deletions also revealed a ∼ 95 kb critical region, which may harbor the GJB2 regulatory element(s).
authors.affiliation: Harvard Medical School Genetics Training Program, Harvard, Cambridge, MA.|Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.|Division of Genomic Diagnostics, The Children's Hospital of Philadelphia, The University of Pennsylvania Perelman School of Medicine, PA.
authors.forename: Ahmad N Abou
authors.initials: AN
authors.lastname: Tayoun
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.|Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.
authors.forename: Heather
authors.initials: H
authors.lastname: Mason-Suares
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.
authors.forename: Ashley L
authors.initials: AL
authors.lastname: Frisella
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.
authors.forename: Mark
authors.initials: M
authors.lastname: Bowser
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Duffy
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Mahanta
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.|Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.
authors.forename: Birgit
authors.initials: B
authors.lastname: Funke
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.|Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.
authors.forename: Heidi L
authors.initials: HL
authors.lastname: Rehm
authors.affiliation: Laboratory for Molecular Medicine, Partners Healthcare Personalized Medicine, Cambridge, MA.|Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.
authors.forename: Sami S
authors.initials: SS
authors.lastname: Amr
mesh_terms: D056905:Chromosome Breakpoints; D000072259:Connexin 26; D000075302:Connexin 30; D017630:Connexins / Q000235:genetics*; D017353:Gene Deletion; D018628:Gene Dosage; D005808:Genes, Recessive; D056426:Genetic Loci; D034381:Hearing Loss / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D017384:Sequence Deletion
chemical_list: D000075302:Connexin 30; D017630:Connexins; C000607042:GJB2 protein, human; C491020:GJB6 protein, human; D000072259:Connexin 26
keywords: DFNB1; GJB2; autosomal-recessive sensorineural hearing loss; copy-number variants; droplet digital PCR