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.