Background

As is known to all, pathogenic bacteria have a serious impact on human health. The development of sensitive, simple, rapid and low-cost bacterial detection method is necessary. Nowadays, some conventional methods (such as plate count, polymerase chain reaction (PCR) and immunological techniques) can not meet the above needs. This work was aimed at providing a new method for addressing these unmet needs.

Result

This study proposed a novel PCR-free and label-free DNA sensor based on multiple linear hybridization chain reaction (ML-HCR) and cloth-based closed bipolar electrochemiluminescence for sensitive and rapid detection of Escherichia coli (E. coli). The target DNA can be obtained from the E. coli genomic DNA by using the restriction enzyme instead of PCR. The auxiliary probe-triggered ML-HCR is carried out with continuous hybridization of two hairpin DNA, and as a result the double stranded DNA is formed to provide a large number of binding sites for Ru(bpy)₃²⁺. The whole detection is PCR-free and label-free, and thus the detection procedure is easier and faster. Under optimized conditions, the linear detection range was from 10² to 10⁷ CFU/mL, and the detection limit was low to 38 CFU/mL. In addition, the proposed DNA sensor has an acceptable selectivity, stability and reproducibility, and is successfully applied to detect E. coli in milk samples with the recoveries from 96.24% to 105.98%.

Significance

The proposed DNA sensor has broad application prospects in the fields of bacterial detection and gene diagnose. Further, this method has potential to be extended for establishing miniaturized, integrated, and automated detection system.

中文翻译：

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一种新型的无 PCR 和无标记布基 DNA 传感器，用于灵敏、快速地检测大肠杆菌

背景

众所周知，致病菌对人体健康有着严重的影响。开发灵敏、简便、快速和低成本的细菌检测方法是必要的。目前，一些常规方法（如平板计数、聚合酶链反应（PCR）和免疫学技术）已不能满足上述需要。这项工作旨在提供一种新方法来解决这些未满足的需求。

结果

本研究提出了一种基于多重线性杂交链式反应 (ML-HCR) 和布基封闭双极电化学发光的新型免 PCR 和免标记 DNA 传感器，用于灵敏、快速检测大肠杆菌( E. coli )。目标 DNA 可以通过使用限制酶而不是 PCR从大肠杆菌基因组 DNA 中获得。_{辅助探针触发的ML-HCR是通过两条发夹DNA连续杂交，形成双链DNA，为Ru(bpy) 3} ²⁺提供大量结合位点. 整个检测过程无需PCR、无需标记，检测过程更加简便快捷。在优化条件下，线性检测范围为10 ²至10 ⁷  CFU/mL，检测限低至38 CFU/mL。此外，所提出的DNA传感器具有可接受的选择性、稳定性和重现性，并成功应用于检测牛奶样品中的大肠杆菌，回收率为96.24%至105.98%。

意义

所提出的DNA传感器在细菌检测和基因诊断领域具有广阔的应用前景。此外，该方法有可能扩展到建立小型化、集成化和自动化的检测系统。