摘要
目的探讨PCR-反向斑点杂交(PCR—RDB)膜芯片技术在遗传性非综合征耳聋患儿基因诊断中的应用价值。方法收集东莞市康复学校38例排除耳聋高危因素的先天性耳聋患儿的外周血2mL,提取基因组DNA,采用自行研究设计的多重PCR扩增结合RDB膜芯片技术针对中国人群4个常见耳聋致病基因20个热点突变进行检测。以Sanger测序技术作为金标准对阳性样本进行验证。结果38例耳聋患儿中,共有16例患儿检出耳聋基因突变,检出率为42.11%,耳聋基因突变的患儿均行家系验证。16例耳聋患儿基因突变中GJB2基因突变6例,检出率为15.79%,其中纯合子3例、杂合子3例;SLC26A4基因突变4例,均为杂合子,检出率为10.53%;MTRNR基因m.1555A>G突变位点2例,检出率为5.26%;复合突变4例,检出率10.53%;未检出GJB3基因突变。16例耳聋基因突变患儿DNA样本经Sanger测序法验证,结果符合率为100%。结论应用针对4个常见耳聋致病基因20个热点突变设计的PCR-RDB膜芯片技术检测先天性耳聋患儿易感基因具有检出率高、快速、准确、经济等特点,是遗传性非综合征耳聋患儿基因筛查的理想方法,具有较好的临床应用前景。
Objective To explore the application value of PCR-reverse dot blot hybridization (PCR-RDB) gene membrane chip technique in genetic diagnosis of hereditary non-syndrome deafness in children.Methods The blood samples(2 mL)were collected from 38 children with congenital deafness,excluding high-risk factors for deaf-ness at Dongguan Rehabilitation School,and then genomic DNA extracted.By using self-designed multiplex-PCR combined with PCR-RDB gene chip technology,20 hot-spot mutations of 4 pathogenic genes of common deafness in Chinese population were detected.Sanger sequencing was used as the gold standard to corroborate the positive samples. Results Among 38 subjects,deafness gene mutations were detected in 16 cases,with a detection rate of 42.11%,and they were all verified by family study.Among 16 cases,6 cases of GJB2 gene mutation(3 cases of homozygote,3 cases of heterozygous),4 cases of SLC26A4 mutation,2 cases of MTRNR (m.1555A>G)mutation,4 cases of compound muta-tion,but none of GJB3 gene mutations.And their detection rates were 15.79%,10.53%,5.26%,10.53%,and 0,re-spectively.DNA samples from 16 children with deafness gene mutation were corroborated by Sanger sequencing,and the compliance rate was 100%.Conclusions For 20 hot-spot mutations of 4 common deafness pathogenic genes,the matc-hing PCR-RDB gene membroine chip technology was designed and the susceptible gene of congenital deafness children was detected.This technique has some advantages like high detection rate,fast,accurate and economical.It is an ideal method for gene screening on hereditary non-syndrome deafness children and has good clinical application prospects.
作者
叶敏南
李文瑞
彭琪
黎四平
彭陆衡
陆小梅
Ye Minnan;Li Wenrui;Peng Qi;Li Siping;Peng Luheng;Lu Xiaomei(Department of Cliniqal Laboratory,Dongguan Eighth People's Hospital (Dongguan Children's Hospital),Dongguan 523325,Guangdong Province,China;Dongguan Institute of Pediatrics,Dongguan 523325,Guangdong Province,China)
出处
《中华实用儿科临床杂志》
CSCD
北大核心
2018年第23期1811-1814,共4页
Chinese Journal of Applied Clinical Pediatrics
基金
广东省科技计划项目(2014A020213001)
东莞市科技计划重点项目(2013108101018).
关键词
遗传性非综合征耳聋
PCR-反向斑点杂交
膜芯片
基因突变
Hereditary nonsyndromes deafness
PCR-reverse dot blot hybridization
Membrane chips
Gene mutation
作者简介
通信作者:陆小梅,Email:lxm020@126.com
