Twenty-five years of quantitative PCR for gene expression analysis.
By H. D. Vanguilder, K. Vrana, W. Freeman. BioTechniques, 2008
1,243 citations recorded by the source corpus.
Following its invention 25 years ago, PCR has been adapted for numerous molecular biology applications. Gene expression analysis by reverse-transcription quantitative PCR (RT-qPCR) has been a key enabling technology of the post-genome era. Since the founding of BioTechniques, this journal has been a resource for the improvements in qPCR technology, experimental design, and data analysis. qPCR and, more specifically, real-time qPCR has become a routine and robust approach for measuring the expression of genes of interest, validating microarray experiments, and monitoring biomarkers. The use of real-time qPCR has nearly supplanted other approaches (e.g., Northern blotting, RNase protection assays). This review examines the current state of qPCR for gene expression analysis now that the method has reached a mature stage of development and implementation. Specifically, the different fluorescent reporter technologies of real-time qPCR are discussed as well as the selection of endogenous controls. The conceptual framework for data analysis methods is also presented to demystify these analysis techniques. The future of qPCR remains bright as the technology becomes more rapid, cost-effective, easier to use, and capable of higher throughput.
Subfields: Molecular Biology
View Paper: https://www.semanticscholar.org/paper/3fb8c8814d17d1563fa3829b82d2fe6eaa59e294
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Citations in the corpus
- PrimerBank: a PCR primer database for quantitative gene expression analysis, 2012 update (2011, background): “Real-time PCR has several major advantages over DNA microarrays, the most important being a large dynamic range, high sensitivity and high specificity (5,6).”
- PrimerBank: a resource of human and mouse PCR primer pairs for gene expression detection and quantification (2009, background): “Hybridization probes are designed to be target specific and can thus minimize nonspecific amplification, but can be difficult to design and costly (5).”
- Recent updates on biogas production - a review (2016, background): “The detected change in fluorescence intensity reflects the concentration of the amplified gene in real time (VanGuilder et al., 2008).”
- Life in soil by the actinorhizal root nodule endophyte Frankia. A review (2010, background): “Instead of measuring dinitrogenase activity in soil, it might be possible to evaluate the specific expression of Frankia nif genes in soil, analysing environmental RNA with appropriate primers and q-PCR (VanGuilder et al. 2008).”
- Primer-BLAST: A tool to design target-specific primers for polymerase chain reaction (2012, background): “This is especially important for real time quantitative PCR (qPCR) where in many cases the amount of PCR product is represented by the total intensity of fluorescence incorporated into amplified DNA and any amplification of unintended targets can affect the measurement [1].”