A Developed Accurate Digital PCR Detection Technology in Food Safety
With the development of the quantitative PCR detection methodologies, different types of PCR techniques have been developed, such as real-time quantitative PCR, competitive real-time PCR, etc. However, all the above techniques only achieve quantitative de
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A Developed Accurate Digital PCR Detection Technology in Food Safety
Abstract With the development of the quantitative PCR detection methodologies, different types of PCR techniques have been developed, such as real-time quantitative PCR, competitive real-time PCR, etc. However, all the above techniques only achieve quantitative detection by standard curves and are considered laborintensive experiments. Based on this background, digital PCR (dPCR), an absolute quantitative detection method, was developed in the 1990s and could achieve quantitative detection without the generation of standard curves. The idea of digital PCR was first raised by Kinzler and Vogelstein in the 1990s based on the limited separation of the original PCR reaction. Digital PCR has the advantage of being highly sensitive, specific, and well stable and having absolute quantitation compared to the original real-time PCR. Over nearly three decades, digital PCR has achieved wide usage in different areas, such as clinical diagnostics, GMO detections, biosensor detections, etc. In this chapter, the background of the development of digital PCR, the concept of digital PCR, and the use of digital PCR in different areas are reviewed in detail. By reading this chapter, the reader could have a better comprehension of all the aspects of digital PCR. Keywords Digital PCR • Absolute quantification • Viral detection • Diagnostic detection • GMO detection
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Background of the Development of Digital PCR
The quantitative detection of all types of samples heavily relies on quantitative polymerase chain reaction (qPCR) to detect and quantify the target sequence. In the past 20 years, based on nucleic acid chemistry, real-time PCR has revolutionized the diagnosis and became the gold standard for the quantification and detection of various samples [1], such as clinical samples, samples of genetically modified organisms, bacterial samples, and others. During quantitative PCR, DNA is amplified until it generates a signal that is embedded in a dye or a sequence-specific fluorescent probe. The period threshold is defined as the number of signals required to achieve the desired level of amplification cycles and the number of DNA molecules for the standard curve based on the presence of the original [2].
© Springer Science+Business Media Singapore 2016 W. Xu, Functional Nucleic Acids Detection in Food Safety, DOI 10.1007/978-981-10-1618-9_7
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A Developed Accurate Digital PCR Detection Technology in Food Safety
Although quantitative PCR has led to significant progress in diagnosis, the technology has obvious limitations. Quantification is based on the standard curve, which requires careful calibration and the same source material. In addition, the signal selection threshold may be adjusted by the operator and is introduced into the main analysis. Due to changes in the structure of the standard curve and potential differences, differences in subjective analysis between commercial kits and standardized protocols can be enormous. Furthermore, even in a tr
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