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Nucl. Acids Res.-2012-Whale-e82.pdf (280.65 kB)

Comparison of microfluidic digital PCR and conventional quantitative PCR for measuring copy number variation

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posted on 2015-07-21, 08:19 authored by Jacqui A. Shaw, A. S. Whale, J. F. Hugget, S. Cowen, V. Speirs, S. Ellison, C. A. Foy, D. J. Scott
One of the benefits of Digital PCR (dPCR) is the potential for unparalleled precision enabling smaller fold change measurements. An example of an assessment that could benefit from such improved precision is the measurement of tumour-associated copy number variation (CNV) in the cell free DNA (cfDNA) fraction of patient blood plasma. To investigate the potential precision of dPCR and compare it with the established technique of quantitative PCR (qPCR), we used breast cancer cell lines to investigate HER2 gene amplification and modelled a range of different CNVs. We showed that, with equal experimental replication, dPCR could measure a smaller CNV than qPCR. As dPCR precision is directly dependent upon both the number of replicate measurements and the template concentration, we also developed a method to assist the design of dPCR experiments for measuring CNV. Using an existing model (based on Poisson and binomial distributions) to derive an expression for the variance inherent in dPCR, we produced a power calculation to define the experimental size required to reliably detect a given fold change at a given template concentration. This work will facilitate any future translation of dPCR to key diagnostic applications, such as cancer diagnostics and analysis of cfDNA.

History

Citation

Nucleic Acids Research, 2012, 40 (11), e82

Author affiliation

/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Medicine/Department of Cancer Studies and Molecular Medicine

Version

  • VoR (Version of Record)

Published in

Nucleic Acids Research

Publisher

Oxford University Press (OUP)

issn

0305-1048

eissn

1362-4962

Acceptance date

2014-02-14

Copyright date

2012

Available date

2015-07-21

Publisher version

http://nar.oxfordjournals.org/content/40/11/e82

Language

en

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