PCR 双向开启硫黄素 T/G 四链体探针无标记检测牛奶中沙门氏菌方法研究

文章提出一种基于聚合酶链式反应(polymerase chain reaction, PCR)的硫黄素T(ThT)/G-四链体探针无标记检测牛奶中沙门氏菌(Salmonella)方法。以沙门氏菌的invA基因为检测目标, 设计特异性发卡引物, 通过PCR和特异功能的G-四链体实现对沙门氏菌的检测。结果表明, 该检测方法操作简便、特异性强、灵敏度高。在最优反应体系下, 沙门氏菌的浓度与信号输出值具有良好的线性关系, 其回归方程为 $ Y=27.149X+0.292, R^{2}=0.981 $, 其线性范围为 $ 10^{2}\sim10^{6} $ CFU/mL。纯菌样品和实际样品都能在细菌浓度为 $ 10^{2} $ CFU/mL时检测出信号, 表明该方法具有抗基质效应。研究结果为食源性致病菌的检测提供了一种新的思路。

A method for the label-free detection of Salmonella in milk by thioflavin T/G-quadruplex probe combined with the polymerase chain reaction(PCR) was established. In this method, the invA gene of Salmonella was employed as the detection target, and the PCR amplification was conducted using specifically designed hairpin primers encoded with G-quadruplex function unit. The results showed that the detection of Salmonella in milk was simple, specific and sensitive. Under the optimal reaction system, there was a good linear relationship between the Salmonella concentration and the signal output value. The regression equation was fitted to be $ Y=27.149X+0.292(R^{2}=0.981) $ with a linear range of $ 10^{2}-10^{6} $ CFU/mL. Moreover, bacteria with concentrations as low as $ 10^{2} $ CFU/mL can be detected in both pure and real samples, indicating that the method has a good anti-matrix effect. This study provides a new avenue for the detection of foodborne pathogens via molecular nucleic acid amplifications.