Calibration of quantitative PCR assays

Quantitative real-time PCR (polymerase chain reaction) assays are increasingly used to measure quantities of nucleic acids in samples. They may be used to provide a high-throughput alternative to more traditional biological assays. In this case, a calibration process may be required to PCR measureme...

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Bibliographic Details
Published in:Journal of agricultural, biological, and environmental statistics biological, and environmental statistics, 2007-09, Vol.12 (3), p.364-378
Main Authors: Roberts, A.M.I, Theobald, C.M, McNeil, M
Format: Article
Language:English
Subjects:
Agronomy. Soil science and plant productions
Bayesian theory
bioassays
Biological and medical sciences
Biometrics, statistics, experimental designs, modeling, agricultural computer applications
Calibration
Censorship
Fundamental and applied biological sciences. Psychology
Fungal diseases
Fungal spores
Generalities. Biometrics, experimentation. Remote sensing
Genetic variation
Linear regression
measurement
model validation
Modeling
Polymerase chain reaction
Product category rules
smut diseases
Spores
statistical models
testing
Tilletia tritici
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Summary:Quantitative real-time PCR (polymerase chain reaction) assays are increasingly used to measure quantities of nucleic acids in samples. They may be used to provide a high-throughput alternative to more traditional biological assays. In this case, a calibration process may be required to PCR measurements onto a more relevant scale. This is most commonly undertaken using simple linear regression. However, such calibration models are usually unrealistic since they ignore the various sources of variation associated with the PCR and conventional assays. Taking account of these various sources is necessary if the errors associated with predictions based on the calibration model are to be well estimated. In this article, we demonstrate a more complete approach to calibration of quantitative PCR. As an example, we develop a Bayesian calibration model for measuring the quantity of the fungus common bunt (Tilletia caries) on wheat seed, based on our understanding of the properties of the assays. As well as illustrating the steps in developing such a model, we show how the fit of the model might be assessed.
ISSN:1085-7117
1537-2693
DOI:10.1198/108571107X227379