NDLI: Accuracy in Copy Number Calling by qPCR and PRT: A Matter of DNA

Content Provider	PubMed Central
Author	Nora, Fernandez-jimenez Castellanos-rubio, Ainara Leticia, Plaza-izurieta Gutierrez, Galder Irastorza, Iñaki Castaño, Luis Vitoria, Juan Carlos Bilbao, Jose Ramon
Editor	Cotterill, Sue
Copyright Year	2011
Abstract	The possible implication of copy number variation (CNV) in the genetic susceptibility to human disease needs to be assessed using robust methods that can be applied at a population scale. In this report, we analyze the performance of the two major techniques, quantitative PCR (qPCR) and paralog ratio test (PRT), and investigate the influence of input DNA amount and template integrity on the reliability of both methods. Analysis of three genes (PRELID1, SYNPO and DEFB4) in a large sample set showed that both methods are prone to false copy number assignments if sufficient attention is not paid to DNA concentration and quality. Accurate normalization of samples is essential for reproducible qPCR because it avoids the effect of differential amplification efficiencies between target and control assays, whereas PRT is generally more sensitive to template degradation due to the fact that longer amplicons are usually needed to optimize sensitivity and specificity of paralog sequence PCR. The use of normalized, high quality genomic DNA yields comparable results with both methods.
Related Links	http://dx.doi.org/10.1371/journal.pone.0028910
Starting Page	28910
File Format	PDF
ISSN	19326203
e-ISSN	19326203
Journal	PLoS ONE
Issue Number	12
Volume Number	6
Language	English
Publisher	Public Library of Science
Publisher Date	2011-12-01
Access Restriction	Open
Rights Holder	Public Library of Science
Subject Keyword	Biochemistry, Genetics and Molecular Biology(all) Agricultural and Biological Sciences(all) Medicine(all) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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