Sample degradation leads to false-positive copy number variation calls in multiplex real-time polymerase chain reaction assays

Anal Biochem. 2009 Mar 15;386(2):288-90. doi: 10.1016/j.ab.2008.11.040. Epub 2008 Dec 7.

Abstract

The recent implication of genomic copy number variations (CNVs) in multiple human genetic disorders has led to increased interest in CNV discovery technologies. There is a growing consensus that, in addition to the method used for detection, at least one additional technology should be employed for validation. Real-time quantitative polymerase chain reaction (qPCR) analysis, incorporating a normal (2N) copy number standard, is commonly used as a means of validating CNVs. Whereas it has previously been reported that formalin-fixed paraffin-embedded (FFPE) DNA samples can yield spurious CNV calls in real-time qPCR assays, here we report that sample degradation under standard laboratory storage conditions generates a significant increase in false-positive CNV results. Results suggest the possibility of biased degradation among genomic regions and emphasize the need to assess sample integrity immediately prior to real-time qPCR experiments.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • False Positive Reactions
  • Gene Dosage
  • Genetic Variation*
  • Genome, Human*
  • Humans
  • Polymerase Chain Reaction*
  • Sensitivity and Specificity