Tang Zhining, Tang Xiaowen, Xue Ling, Guan Minxin
School of Laboratory Medicine and Life Sciences, Attardi Institute of Mitochondrial Biomedicine, Wenzhou Medical University, Wenzhou 325035, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2021 Jan 30;41(1):151-156. doi: 10.12122/j.issn.1673-4254.2021.01.23.
To explore the feasibility of detecting maternal hereditary mitochondrial tRNA15927G>A (m.15927G>A) mutation using buccal swabs.
We performed sequence analysis of mitochondrial DNA in blood samples from 2070 cases of maternal hereditary mitochondrial disease in the First Affiliated Hospital of Wenzhou Medical University, and identified 3 patients with m.15927G>A mutation.Buccal swabs and blood samples were obtained from the 3 patients (mutation group) and 3 normal volunteers (control group).After extracting whole genomic DNA from all the samples, the DNA concentration and purity were analyzed.The PCR products were subjected to dot blot hybridization, Southern blot hybridization, and DNA sequencing analysis to verify the feasibility of detecting m.15927G>A mutation using buccal swabs.
There was no significant difference in DNA concentration extracted from buccal swabs and blood samples in either the mutation group or the control group ( > 0.05), but the purity of manually extracted oral mucosa DNA was significantly lower than that of whole blood and oral mucosa DNA extracted using commercial kits ( < 0.05).Dot blot hybridization and Southern blot hybridization both yielded positive results in the control group but negative results in the mutation group.DNA sequencing identified m.15927G>A mutation in all the samples from the mutation group.
Buccal swabs collection accurate is an accurate and sensitive method for the detection of m.15927G>A mutation.
探讨使用口腔拭子检测母体遗传性线粒体tRNA15927G>A（m.15927G>A）突变的可行性。
我们对温州医科大学附属第一医院2070例母体遗传性线粒体疾病患者的血液样本进行线粒体DNA序列分析，鉴定出3例携带m.15927G>A突变的患者。从这3例患者（突变组）和3名正常志愿者（对照组）采集口腔拭子和血液样本。从所有样本中提取全基因组DNA后，分析DNA浓度和纯度。对PCR产物进行斑点杂交、Southern杂交和DNA测序分析，以验证使用口腔拭子检测m.15927G>A突变的可行性。
突变组和对照组中，从口腔拭子和血液样本中提取的DNA浓度均无显著差异（>0.05），但手工提取的口腔黏膜DNA纯度显著低于全血和使用商业试剂盒提取的口腔黏膜DNA（<0.05）。斑点杂交和Southern杂交在对照组中均得到阳性结果，而在突变组中得到阴性结果。DNA测序在突变组的所有样本中均鉴定出m.15927G>A突变。
口腔拭子采集是检测m.15927G>A突变的一种准确且灵敏的方法。
