运用实时定量聚合酶链反应对中国某聋哑人群中线粒体DNA C1494T突变进行快速筛查。
Rapid screening for the mitochondrial DNA C1494T mutation in a deaf population in China using real-time quantitative PCR.
作者信息
Li Qi, Yuan Yong-Yi, Huang De-Liang, Han Dong-Yi, Dai Pu
机构信息
Department of Otolaryngology, Nanjing Children's Hospital, Nanjing Medical University, Jiangsu, China.
出版信息
Acta Otolaryngol. 2012 Aug;132(8):814-8. doi: 10.3109/00016489.2012.664781. Epub 2012 Apr 12.
CONCLUSION
Real-time quantitative polymerase chain reaction (qPCR) with a TaqMan minor groove binding (MGB) probe is useful for large-scale screening for the C1494T mutation. The mitochondrial DNA(mtDNA) C1494T mutation has a low carrier frequency in Chinese patients with nonsyndromic hearing loss.
OBJECTIVE
To develop a simple, rapid, and reliable real-time qPCR assay based on TaqMan technology using a new MGB probe for detecting the mtDNA C1494T mutation directly, and to investigate the carrier frequency in nonsyndromic deaf Chinese subjects.
METHODS
A TaqMan-MGB probe was constructed. Peripheral blood samples were collected from 3133 nonsyndromic deaf patients and genomic DNA was extracted. A real-time qPCR using MGB probes (wild-type) in a single tube was used to detect the mtDNA C1494T mutation. The results were then compared to the DNA sequence of the PCR products.
RESULTS
A total of 13 of 3133 (0.4%) Chinese nonsyndromic hearing loss patients were C1494T-positive. The results of the TaqMan-MGB probe method were consistent with those of sequencing.
结论
采用TaqMan小沟结合（MGB）探针的实时定量聚合酶链反应（qPCR）技术有助于大规模筛查C1494T突变。线粒体DNA（mtDNA）C1494T突变在中国非综合征性听力损失患者中的携带频率较低。
目的
基于TaqMan技术开发一种简单、快速且可靠的实时qPCR检测方法，使用新型MGB探针直接检测mtDNA C1494T突变，并调查中国非综合征性耳聋患者中的携带频率。
方法
构建TaqMan-MGB探针。采集3133例非综合征性耳聋患者的外周血样本并提取基因组DNA。使用单管中的MGB探针（野生型）进行实时qPCR以检测mtDNA C1494T突变。然后将结果与PCR产物的DNA序列进行比较。
结果
3133例中国非综合征性听力损失患者中共有13例（0.4%）C1494T呈阳性。TaqMan-MGB探针法的结果与测序结果一致。
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