A novel emulsion PCR method coupled with fluorescence spectrophotometry for simultaneous qualitative, quantitative and high-throughput detection of multiple DNA targets

文献类型： 外文期刊

第一作者： Zhao, Xiao

作者： Zhao, Xiao;Zhao, Kai;Zhao, Xiao;Zhao, Kai;Du, Yanan;Xu, Yan;Shi, Wei;Ren, Fangfang;Wu, Yangyang;Hu, Ruili;Fan, Xiaorui;Zhang, Qi;Zhang, Xiaoxia;Zhang, Wanjing;Wu, Wenjing;Zhao, Binan;Shi, Bin;Zhao, Huanzhen

作者机构：

期刊名称：SCIENTIFIC REPORTS （ 影响因子：4.379； 五年影响因子：5.133 ）

ISSN： 2045-2322

年卷期： 2019 年 9 卷

页码：

收录情况： SCI

摘要： We constructed and validated a novel emulsion PCR method combined with fluorescence spectrophotometry (EPFS) for simultaneous qualitative, quantitative and high-throughput detection of multiple DNA targets. In a single reaction set, each pair of primers was labeled with a specific fluorophore. Through emulsion PCR, a target DNA was amplified in droplets that functioned as micro-reactors. After product purification, different fluorescent-labeled DNA products were qualitatively analyzed by the fluorescent intensity determination. The sensitivity and specificity of the system was examined using four kinds of genetically modified (GM) maize. The qualitative results revealed high specificity and sensitivity of 0.5% (w/w). In addition, the quantitative results revealed that the absolute limit of detection was 10(3) copies, showing good repeatability. Moreover, the reproducibility assays were further performed using four foodborne pathogenic bacteria to further evaluate the applicability of the system. Consequently, the same qualitative, quantitative and high-throughput results were confirmed with the four GM maize. To sum up, the new EPFS system is the first analytical technology of this kind that enables simultaneous qualitative, quantitative and high-throughput analysis of multiple genes.

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