Analysis of rhodopsin gene in patients with retinitis pigmentosa using allele-specific polymerase chain reaction.

Point mutations within the rhodopsin gene have been found recently in some patients with autosomal dominant retinitis pigmentosa (ADRP). Currently, four types of point mutations at codons 23, 58 and 347 have been identified. The purposes of this study were to establish simple methods for screening patients with retinitis pigmentosa (RP) to detect these point mutations, and to apply these methods to determine if these mutations are found in Japanese patients with RP. Utilizing the polymerase chain reaction (PCR), a one-step method was developed to detect point mutations at codon 23. This method was then applied to screen genomic DNAs from 30 patients with various types of RP, including ADRP, autosomal recessive RP, simplex RP, Leber's congenital amaurosis or Usher's syndrome. Subsequently, point mutations at codons 58 and 347 were detected by restriction enzyme digestion (Dde I or Msp I) of exons 1 and 5 amplified by PCR. To date, no mutations have been found in codons 23 and 58 in Japanese patients. By using the allele-specific PCR, however, two patients from one pedigree of ADRP were confirmed to have a C-to-T transition at the second nucleotide of codon 347, which results in the substitution of leucine for proline. Our findings indicated the availability of this simple method for detecting these point mutations.