RESUMEN
We have investigated the prevalence of mutations in the connexin 26 (GJB2) gene in Omani population using both PCR-RFLP and direct DNA sequencing methods. Two common GJB2 gene mutations (35delG and 167delT) were screened in 280 healthy controls and 95 deaf patients using two different PCR-RFLP methods. To investigate other GJB2 mutations, we have amplified and sequenced DNA from 51 unrelated deaf patients and 17 control subjects. None of the samples studied, either by RFLP or sequencing, revealed any deafness-associated mutations in the coding region of the GJB2 gene. These findings disagree with many reports on the GJB2 gene, describing various mutations as the cause of congenital recessive deafness. Although, an amino acid substitution (S86T) was identified by sequencing, we conclude that this change could not be associated with deafness since it was present in all the control and patient samples sequenced.
Asunto(s)
Conexinas/genética , Sordera/genética , Secuencia de Aminoácidos , Secuencia de Bases , Conexina 26 , ADN/química , ADN/genética , Análisis Mutacional de ADN , Humanos , Mutación , Omán , Polimorfismo de Longitud del Fragmento de RestricciónRESUMEN
BACKGROUND: Several mutations described in the connexin-26 gene cause nonsyndromic autosomal recessive deafness (NARD). The prevalence of two frame-shift mutations, known as 35delG and 167delT, was relatively high in patients with NARD from different populations. METHODS AND RESULTS: A seminested PCR test has been developed for simultaneous detection of two common mutations in the connexin-26 gene. The test is based on PCR amplification of a 285-bp DNA fragment that covers both the 35delG and 167delT mutations. The latter mutation destroys a Pst I site and is easily detected by Pst I digestion of the 285-bp DNA fragment. However, the 35delG mutation does not destroy or create a restriction site. To create a site, we designed a mismatched primer that generated an EcoN I site in an 87-bp DNA fragment, but only if the 35delG mutation was present. The test was validated using five DNA samples previously characterized for the connexin-26 mutations. After validation, we screened 45 unrelated patients with NARD and 280 healthy Omani subjects for the presence or absence of the 35delG and 167delT mutations. Neither mutation was found to be present in patients or control subjects. CONCLUSION: We developed a seminested PCR test for the simultaneous detection of both common mutations in the connexin-26 gene. In our analysis of 45 patients and 280 control subjects, the 35delG and 167delT mutations were absent in both groups.