Alexandra Bogožalec Košir (Author), Sabine Muller (Author), Jana Žel (Author), Mojca Milavec (Author), Allison C. Mallory (Author), David Dobnik (Author)

Abstract

The proliferation of genetically modified organisms (GMOs) presents challenges to GMO testing laboratories and policymakers. Traditional methods, like quantitative real-time PCR (qPCR), face limitations in quantifying the increasing number of GMOs in a single sample. Digital PCR (dPCR), specifically multiplexing, offers a solution by enabling simultaneous quantification of multiple GMO targets. This study explores the use of the Naica six-color Crystal dPCR platform for quantifying five GM soybean lines within a single six-plex assay. Two four-color assays were also developed for added flexibility. These assays demonstrated high specificity, sensitivity (limit of detection or LOD < 25 copies per reaction) and precision (bias to an estimated copy number concentration <15%). Additionally, two approaches for the optimization of data analysis were implemented. By applying a limit-of-blank (LOB) correction, the limit of quantification (LOQ) and LOD could be more precisely determined. Pooling of reactions additionally lowered the LOD, with a two- to eight-fold increase in sensitivity. Real-life samples from routine testing were used to confirm the assays’ applicability for quantifying GM soybean lines in complex samples. This study showcases the potential of the six-color Crystal dPCR platform to revolutionize GMO testing, facilitating comprehensive analysis of GMOs in complex samples.

Keywords

digitalna PCR;digitalna verižna reakcija s polimerazo;kvantifikacija;multipleksiranje;gensko spremenjeni organizmi;6-barvni sistem;diagnostika virusov;virologija;digital PCR;dPCR;quantification;multiplexing;genetically modified organisms;6-color system;virus diagnostics;virology;

Data

Language: English
Year of publishing:
Typology: 1.01 - Original Scientific Article
Organization: NIB - National Institute of Biology
UDC: 578
COBISS: 173360131 Link will open in a new window
ISSN: 2304-8158
Views: 166
Downloads: 65
Average score: 0 (0 votes)
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Other data

Secondary language: Slovenian
Secondary keywords: digitalna PCR;digitalna verižna reakcija s polimerazo;kvantifikacija;multipleksiranje;gensko spremenjeni organizmi;6-barvni sistem;diagnostika virusov;virologija;
Source comment: Soavtorji: Sabine Muller, Jana Žel, Mojca Milavec, Allison C. Mallory and David Dobnik; Nasl. z nasl. zaslona; Opis vira z dne 23. 11. 2023;
Pages: str. [1]-17
Volume: ǂiss. ǂ22
Issue: ǂart. no. ǂ4156
Chronology: Nov. 2023
DOI: 10.3390/foods12224156
ID: 23284779