Magnetic Bead Chain-Based Continuous-Flow DNA Extraction for Microfluidic PCR Detection of Salmonella.

Yuhe Wang,Yuanjie Liu,Wuzhen Qi,Lei Wang,Jianhan Lin

doi:10.3390/mi12040384

Abstract

Nucleic acid extraction is crucial for PCR detection of pathogenic bacteria to ensure food safety. In this study, a new magnetic extraction method was developed using 3D printing and magnetic silica beads (MSBs) to extract the target DNA from a large volume of bacterial sample and combined with microfluidic PCR to determine the bacteria. After proteinase K was added into a bacterial sample to lyse the bacteria and release the DNA, it was continuous-flow injected into the serpentine channel of the extraction chip, where magnetic silica bead chains had been formed in advance using a homogeneous magnetic field generated by two concentric semicircle magnets to capture the MSBs. Then, the flowing DNA was captured by the MSB chains, washed with alcohol, dried with gas, and eluted with deionized water to obtain the purified and concentrated DNA. Finally, the extracted DNA templates were injected into a microfluidic PCR chip with lyophilized amplification reagents and determined using a commercial qPCR device. The experimental results showed that the DNA extraction efficiency was more than 90%, and the lower detection limit of Salmonella was 102 CFU/mL. This new Salmonella detection method is promising to provide the rapid, sensitive, and simultaneous detection of multiple foodborne pathogens.

Highlights

Foodborne illness is an important public health issue, and an estimated 600 million people fall ill after eating contaminated foods every year, causing 420,000 deaths according to the statistics of World Health Organization [1]
We developed a new continuous-flow DNA extraction method and combined it with the microfluidic qPCR method for the rapid detection of Salmonella
The distribution of the homogenous magnetic field in the serpentine channel is the key to forming the magnetic silica beads (MSBs) chains and has a great impact on the DNA extraction efficiency

Summary

Introduction

Foodborne illness is an important public health issue, and an estimated 600 million people fall ill after eating contaminated foods every year, causing 420,000 deaths according to the statistics of World Health Organization [1]. Salmonella is one of the leading causes for foodborne diseases [2], and the United States Department of Agriculture (USDA) claimed that the annual cost due to the infection of Salmonella in the US was about 4.1 billion dollars [3]. Rapid screening of Salmonella is critical to prevent the spread of foodborne diseases. Many rapid methods have been developed to detect foodborne bacteria. With the advance of PCR technology, different PCR methods have been developed for the rapid detection of foodborne pathogens, such as qPCR [7,8], multiplex

Methods

Results

Conclusion