title: Design and Validation of a New MLPA-Based Assay for the Detection of RS1 Gene Deletions and Application in a Large Family with X-Linked Juvenile Retinoschisis.
abstract: OBJECTIVE
  X-linked juvenile retinoschisis (XLRS) is a severe ocular disorder that can evolve to blindness. More than 200 different disease-causing mutations have been reported in the RS1 gene and approximately 10% of these are deletions. Since transmission is X-linked, males are always affected and females are usually carriers. The identification of female carriers is always important and poses a technical challenge. Therefore, we sought to develop a multiplex ligation dependent probe amplification (MLPA)-based method to identify deletions or duplications in this gene. We then used our assay to study a large XLRS family.
  
  
  METHODS
  We designed six probes specific for each RS1 exon and then optimized and validated our method using control samples with known gene deletions. In the XLRS family, RS1 gene copy number variation was assessed by "home-made" MLPA analysis and by single nucleotide polymorphism (SNP) array analysis using the CytoScan HD Array. Direct sequencing was used for deletion breakpoint mapping.
  
  
  RESULTS
  Our assay detected all deletions in control samples. All affected males of the family were positive for a deletion of exon 2 of the RS1 gene (RS1:NM_000330:c.53-?_78+?del). Carrier females were also identified.
  
  
  CONCLUSIONS
  Our method is easily replicated, reliable, and inexpensive and allows female carriers to be detected. This is the first report of deep characterization of a whole exon deletion in the RS1 gene.
authors.affiliation: 1 MAGI Non-Profit Human Medical Genetics Institute , Rovereto, Trento, Italy .|2 Department of Medical and Surgical Sciences (DIMEC), University of Bologna , Bologna, Italy .
authors.forename: Annalisa
authors.initials: A
authors.lastname: Nicoletti
authors.affiliation: 3 "G.B. Bietti" Foundation-IRCCS, Roma, Italy .
authors.forename: Lucia
authors.initials: L
authors.lastname: Ziccardi
authors.affiliation: 1 MAGI Non-Profit Human Medical Genetics Institute , Rovereto, Trento, Italy .
authors.forename: Paolo Enrico
authors.initials: PE
authors.lastname: Maltese
authors.affiliation: 1 MAGI Non-Profit Human Medical Genetics Institute , Rovereto, Trento, Italy .
authors.forename: Sabrina
authors.initials: S
authors.lastname: Benedetti
authors.affiliation: 4 Medical Genetics Service, "IRCCS Casa Sollievo della Sofferenza," San Giovanni Rotondo, Italy .
authors.forename: Orazio
authors.initials: O
authors.lastname: Palumbo
authors.affiliation: 4 Medical Genetics Service, "IRCCS Casa Sollievo della Sofferenza," San Giovanni Rotondo, Italy .
authors.forename: Michelina
authors.initials: M
authors.lastname: Rendina
authors.affiliation: 4 Medical Genetics Service, "IRCCS Casa Sollievo della Sofferenza," San Giovanni Rotondo, Italy .
authors.forename: Leonardo
authors.initials: L
authors.lastname: D'Agruma
authors.affiliation: 5 Institute of Ophthalmology, Policlinico Gemelli, Catholic University , Roma, Italy .
authors.forename: Benedetto
authors.initials: B
authors.lastname: Falsini
authors.affiliation: 6 DNA Diagnostic Laboratory, Ophthalmic Genetics and Visual Function Branch, National Eye Institute/NIH , Bethesda, Maryland.
authors.forename: Xinjing
authors.initials: X
authors.lastname: Wang
authors.affiliation: 1 MAGI Non-Profit Human Medical Genetics Institute , Rovereto, Trento, Italy .
authors.forename: Matteo
authors.initials: M
authors.lastname: Bertelli
mesh_terms: D000328:Adult; D056915:DNA Copy Number Variations; D015342:DNA Probes; D005091:Exons; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D005260:Female; D017353:Gene Deletion; D006580:Genetic Carrier Screening / Q000379:methods; D006801:Humans; D008297:Male; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods; D010375:Pedigree; D015203:Reproducibility of Results; D041441:Retinoschisis / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion
chemical_list: D015342:DNA Probes; D005136:Eye Proteins; C108726:RS1 protein, human
keywords: MLPA; RS1; XLRS; retinoschisis