David Dobnik (Author), Tina Demšar (Author), Ingrid Huber (Author), Lars Gerdes (Author), Sylvia Broeders (Author), Nancy Roosens (Author), Frédéric Debode (Author), Gilbert Berben (Author), Jana Žel (Author)

Abstract

Digital PCR (dPCR), as a new technology in the field of genetically modified (GM) organism (GMO) testing, enables determination of absolute target copy numbers. The purpose of our study was to test the transferability of methods designed for quantitative PCR (qPCR) to dPCR and to carry out an inter-laboratory comparison of the performance of two different dPCR platforms when determining the absolute GM copy numbers and GM copy number ratio in reference materials certified for GM content in mass fraction. Overall results in terms of measured GM% were within acceptable variation limits for both tested dPCR systems. However, the determined absolute copy numbers for individual genes or events showed higher variability between laboratories in one third of the cases, most possibly due to variability in the technical work, droplet size variability, and analysis of the raw data. GMO quantification with dPCR and qPCR was comparable. As methods originally designed for qPCR performed well in dPCR systems, already validated qPCR assays can most generally be used for dPCR technology with the purpose of GMO detection.

Keywords

digital PCR;droplet digital PCR;absolute quantification;reference materials;GMO quantification;

Data

Language: English
Year of publishing:
Typology: 1.01 - Original Scientific Article
Organization: NIB - National Institute of Biology
UDC: 604.6
COBISS: 4500303 Link will open in a new window
ISSN: 1618-2642
Views: 16
Downloads: 6
Average score: 0 (0 votes)
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Other data

Pages: str. 211-221
Volume: ǂVol. ǂ410
Issue: ǂiss. ǂ1
Chronology: 2018
DOI: 10.1007/s00216-017-0711-1
ID: 24586341