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This Article Full Text Full Text (PDF) All Versions of this Article: iovs.07-1656v1 49/9/3799    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Jin, Z.-B. Articles by Takahashi, M. PubMed PubMed Citation Articles by Jin, Z.-B. Articles by Takahashi, M.

Allelic Copy Number Variation in FSCN2 Detected Using Allele-Specific Genotyping and Multiplex Real-Time PCRs

¹From the Laboratory for Retinal Regeneration, RIKEN Center for Developmental Biology, Kobe, Hyogo, Japan; and the ²Department of Ophthalmology and Visual Science, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

PURPOSE. Allelic copy number variation (CNV) may alter the functional effects of a heterozygous mutation. The underlying mechanisms and their roles in hereditary diseases, however, are largely unknown. In the present study an FSCN2 mutation was examined that has been reported, not only in patients with retinitis pigmentosa (RP), but also in the normal population.

METHODS. Experiments were performed to investigate the gene and allele copy numbers of FSCN2 in patients with RP who have the c.72delG mutation as well as healthy subjects with or without the mutation. A real-time PCR-based genotyping approach was established that used a real-time PCR assay to qualify the copy numbers of both the wild-type and mutant alleles of the FSCN2 gene.

RESULTS. Three patients with RP and three normal subjects had an equal ratio of the alleles. Of interest, another patient had an asymmetric allele ratio (4:1) of the copy number of the wild-type allele, compared with that of the mutant allele. These findings were further verified using quantitative assays. An allele-specific methylation assay demonstrated a random methylation pattern in the FSCN2 gene.

CONCLUSIONS. The copy numbers of the FSNC2 gene and of each allele in the mutant samples were quantified. The findings excluded the possibility that allelic CNV was associated with RP, suggesting that the c.72delG variant is not the primary cause of RP. It is not likely that the FSCN2 gene is imprinted differentially. The real-time PCR-based genotyping method developed in this study is useful for investigations of allelic asymmetries within genomic regions with CNVs.