Rapid and reliable detection of nonsyndromic hearing loss mutations by multicolor melting curve analysis

Abstract

Hearing loss is a common birth defect worldwide. The GJB2, SLC26A4, MT-RNR1 and MT-TS1 genes have been reported as major pathogenic genes in nonsyndromic hearing loss. Early genetic screening is recommended to minimize the incidence of hearing loss. We hereby described a multicolor melting curve analysis (MMCA)-based assay for simultaneous detection of 12 prevalent nonsyndromic hearing loss-related mutations. The three-reaction assay could process 30 samples within 2.5 h in a single run on a 96-well thermocycler. Allelic types of each mutation could be reproducibly obtained from 10 pg ∼100 ng genomic DNA per reaction. For the mitochondrial mutations, 10% ∼ 20% heteroplasmic mutations could be detected. A comparison study using 501 clinical samples showed that the MMCA assay had 100% concordance with both SNaPshot minisequencing and Sanger sequencing. We concluded that the MMCA assay is a rapid, convenient and cost-effective method for detecting the common mutations, and can be expectedly a reliable tool in preliminary screening of nonsyndromic hearing loss in the Chinese Han population.