Establishment of a Gene Detection System for Hotspot Mutations of Hearing Loss

Abstract

Hearing loss is an etiologically heterogeneous trait with a high incidence in China. Though conventional newborn hearing screening program has been widely adopted, gene detection can significantly improve the means of early discovering genetic risk factors. Thus, simple and efficient methods with higher sensitivity and lower cost for detecting hotspot mutations of hearing loss are urgently requested. Here we established a mutation detection system based on multiple fluorescent probe technique, which can detect and genotype nine hotspot mutations of four prominent hearing loss-related genes in two reactions on a four-channel real-time PCR instrument, including GJB2 (rs750188782, rs80338943, rs1110333204, and rs80338939), GJB3 (rs74315319), SLC26A4 (rs111033313 and rs121908362), and mtDNA 12S rRNA (rs267606617 and rs267606619). This system is with high sensitivity that enables detecting as low as 10 DNA copies samples per reaction. A comparison study in 268 clinical samples showed that the detection system had 100% concordance to Sanger sequencing. Besides, blood and saliva samples can be directly detected without DNA extraction process, which greatly simplifies the manipulation. The new system with high sensitivity, accuracy, and specimen type compatibility can be expectedly a reliable tool in clinical application.