Abstract
During the last few years, several innovative technologies have become available for performing sensitive and accurate genetic analyses. These techniques use fluorescent detection strategies in combination with nucleic acid amplification protocols. Most commonly used is the real-time polymerase chain reaction (PCR). To achieve the maximum potential of a realtime PCR assay, several parameters must be evaluated and optimized independently. This chapter describes the different steps necessary for establishing a molecular beacon real-time PCR assay: (1) target design, (2) primer design, (3) optimization of the amplification reaction conditions using SYBR Green, (4) molecular beacon design, and (5) molecular beacon synthesis and characterization. The last section provides an example of a multiplex quantitative real-time PCR.
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Vet, J.A.M., Marras, S.A.E. (2005). Design and Optimization of Molecular Beacon Real-Time Polymerase Chain Reaction Assays. In: Herdewijn, P. (eds) Oligonucleotide Synthesis. Methods in Molecular Biology, vol 288. Humana Press. https://doi.org/10.1385/1-59259-823-4:273
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DOI: https://doi.org/10.1385/1-59259-823-4:273
Publisher Name: Humana Press
Print ISBN: 978-1-58829-233-9
Online ISBN: 978-1-59259-823-6
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