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Multiplex Droplet Digital PCR Protocols for Quantification of GM Maize Events

Multiplex Droplet Digital PCR Protocols for Quantification of GM Maize Events

David Dobnik, Bjørn Spilsberg, Alexandra Bogožalec Košir, Dejan Štebih, Dany Morisset, Arne Holst-Jensen, Jana Žel

Methods Mol Biol. 2018;1768:69-98.

PMID: 29717438

Abstract:

The standard-curve based simplex quantitative polymerase chain reaction (qPCR) has been the gold standard for DNA target quantification for more than a decade. The large and growing number of individual analyses needed to test for genetically modified organisms (GMOs) is reducing the cost-effectiveness of qPCR. Droplet digital PCR (ddPCR) enables absolute quantification without standard curves, avoids the amplification efficiency bias observed with qPCR, allows more accurate estimations at low target copy numbers and, in combination with multiplexing, significantly improves cost efficiency. Here we describe two protocols for multiplex quantification of GM maize events: (1) nondiscriminating, with multiplex quantification of targets as a group (12 GM maize lines) and (2) discriminating, with multiplex quantification of individual targets (events). The first enables the quantification of twelve European Union authorized GM maize events as a group with only two assays, but does not permit determination of the individual events present. The second protocol enables the quantification of four individual targets (three GM events and one endogene) in a single reaction. Both protocols can be modified for quantification of any other DNA target.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AS24551	Maize GMO Standard ERM-BF423			Price

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