title: Rapid detection of SAG 926delA mutation using real-time polymerase chain reaction.
abstract: OBJECTIVE
  Mutation 926delA of the arrestin/S-antigen SAG gene is the main cause of Oguchi disease in the Japanese. The purpose of this study was to develop a rapid diagnostic assay to detect mutations in the SAG gene.
  
  
  METHODS
  Two sequence-specific primers and fluorophore-labeled probes for exon 11 of the SAG gene were designed, and the region spanning the mutations was amplified by polymerase chain reaction (PCR) using the LightCycler detection system (Roche Diagnostics, Mannheim, Germany). The mutations were then identified by melting curve analyses of the hybrid formed between the PCR product and a specific fluorescent probe.
  
  
  RESULTS
  We clearly distinguished each SAG genotype (homozygous and heterozygous 926delA and wild type) by the distinct melting peaks at different temperatures. One thermal cycling required approximately 54 min to process, and the results were 100% in concordance with the genotypes determined by DNA sequencing.
  
  
  CONCLUSIONS
  We have succeeded in developing a rapid method to detect the most frequent mutation in the SAG gene. This method will help in identifying gene mutations associated with Oguchi disease with a rapid and reliable identification or the exclusion of the frequent mutations in the SAG gene.
authors.affiliation: Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan. yosida@eye.med.kyushu-u.ac.jp
authors.forename: Shigeo
authors.initials: S
authors.lastname: Yoshida
authors.forename: Yoko
authors.initials: Y
authors.lastname: Yamaji
authors.forename: Ayako
authors.initials: A
authors.lastname: Yoshida
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Ikeda
authors.forename: Ken
authors.initials: K
authors.lastname: Yamamoto
authors.forename: Tatsuro
authors.initials: T
authors.lastname: Ishibashi
mesh_terms: D000293:Adolescent; D019393:Arrestin / Q000032:analysis* / Q000235:genetics*; D003623:Dark Adaptation; D004596:Electroretinography; D005654:Fundus Oculi; D017353:Gene Deletion; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D008297:Male; D009154:Mutation; D009216:Myopia / Q000150:complications; D009755:Night Blindness / Q000150:complications / Q000175:diagnosis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013696:Temperature; D013997:Time Factors
chemical_list: D019393:Arrestin