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Genet Test Mol Biomarkers. 2017 Feb;21(2):116-121. doi: 10.1089/gtmb.2016.0257. Epub 2016 Dec 20.

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.

Author information

1
1 MAGI Non-Profit Human Medical Genetics Institute , Rovereto, Trento, Italy .
2
2 Department of Medical and Surgical Sciences (DIMEC), University of Bologna , Bologna, Italy .
3
3 "G.B. Bietti" Foundation-IRCCS, Roma, Italy .
4
4 Medical Genetics Service, "IRCCS Casa Sollievo della Sofferenza," San Giovanni Rotondo, Italy .
5
5 Institute of Ophthalmology, Policlinico Gemelli, Catholic University , Roma, Italy .
6
6 DNA Diagnostic Laboratory, Ophthalmic Genetics and Visual Function Branch, National Eye Institute/NIH , Bethesda, Maryland.

Abstract

AIMS:

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.

CONCLUSION:

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.

KEYWORDS:

MLPA; RS1; XLRS; retinoschisis

PMID:
27997221
PMCID:
PMC5335781
DOI:
10.1089/gtmb.2016.0257
[Indexed for MEDLINE]
Free PMC Article

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