title: A PCR-RFLP test for simultaneous detection of two single-nucleotide insertions in the Connexin-26 gene promoter.
abstract: Comparisons of Connexin-26 (GJB2) gene sequences available in the GenBank data base indicate the presence of a polymorphism in the promoter, but no easy method is available for the detection of this polymorphism. We have developed a PCR-RFLP test for simultaneous detection of two single nucleotide insertions (G and A) in the GJB2 promoter. The test is based on amplification of a 146-bp DNA fragment, which was digested with Mae I to detect the G insertion in the promoter. A similar digestion with Hinf I detects the A insertion. The test was validated using direct DNA sequencing of amplified DNA from 33 samples. After validation, we have used it to investigate DNA samples from 160 control subjects and 51 unrelated patients with nonsyndromic autosomal recessive deafness. All of the samples analyzed using the PCR test and DNA sequencing were found to contain both the G and A insertions in the GJB2 gene promoter. This PCR test will be useful in studying the prevalence of these two insertions in other populations.
authors.affiliation: Biochemistry Department, Sultan Qaboos University, College of Medicine, Postal Code 123 Muscat, Sultanate of Oman. mssimsek@omantel.tel.om
authors.forename: Mehmet
authors.initials: M
authors.lastname: Simsek
authors.forename: Nadia
authors.initials: N
authors.lastname: Al-Wardy
authors.forename: Aisha
authors.initials: A
authors.lastname: Al-Khayat
authors.forename: Mazin
authors.initials: M
authors.lastname: Al-Khabory
mesh_terms: D000072259:Connexin 26; D017630:Connexins / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods; D003638:Deafness / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic
chemical_list: D017630:Connexins; C000607042:GJB2 protein, human; D000072259:Connexin 26