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Real-time PCR methods for the detection of DNA constructs with the nptII gene for the detection of genetically modified plants in food, feed and seed

  • Ralf Reiting
GENERELLE ASPEKTE

Abstract

For routine analysis of the most different food and feed matrices for genetical modification screening methods have increasingly been applied during the past years as the first step of detection. Screening for frequently used regulation elements and recombinant DNA-constructs by the use of real-time PCR is the state of the art. By combining several screening methods, the number of lines of genetically modified plants existing in one sample can be delimited. Apart from the genetically modified plants authorized in the EC, for which reference material and detection methods have to be made available by the placer on the market, there has been an increase of unauthorized GMO on the European market recently, for which event specific detection methods are not available as a rule. It does therefore have to be the target of routine controls by the laboratories charged with surveillance to be able to detect as much as possible of genetically modified plants with the aid of just a few screening PCR methods. Three different construct specific real-time PCR methods are presented in this study. They amplify the junction sequence from the 35S-promoter or the nos-promoter to the nptII gene. Furthermore, a gene specific real-time PCR method for the detection of DNA sequences of the nos-promoter from Agrobacterium tumefaciens is presented.

Keywords

PCR Detection of DNA constructs Genetically modified plants nptII Food Feed Seed 

Real-Time PCR-Methoden für den Nachweis von DNA-Konstrukten mit dem nptII-Gen zum Nachweis gentechnisch veränderter Pflanzen in Lebensmitteln, Futtermitteln und Saatgut

Zusammenfassung

Zur Routine-Untersuchung verschiedener Lebens- und Futtermittelmatrizes auf gentechnische Veränderungen werden in den letzten Jahren zunehmend Screeningverfahren als erster Nachweisschritt angewendet, mit denen häufig eingesetzte Regulationselemente und gentechnische Konstrukte nachgewiesen werden. Der Stand der Technik ist hier die Anwendung der real-time PCR. Durch Kombination mehrerer Screeningverfahren kann die Zahl der in einer Probe vorhandenen Linien gentechnisch veränderter Pflanzen eingegrenzt werden. Neben den in der europäischen Union zugelassenen gentechnisch veränderten Pflanzen, für die Referenzmaterial und Nachweisverfahren durch den Inverkehrbringer zur Verfügung gestellt werden müssen, gelangten in jüngster Vergangenheit vermehrt nicht zugelassene GVO auf den europäischen Markt, für die in der Regel keine Event-spezifischen Nachweisverfahren zur Verfügung stehen. Ziel der Routine-Kontrollen der für die Überwachung zuständigen Labore muss es daher sein, mit Hilfe weniger Screening-PCR-Verfahren eine möglichst große Anzahl an gentechnisch veränderten Pflanzen detektieren zu können. Hier werden drei verschiedene Konstrukt-spezifische real-time PCR-Verfahren vorgestellt, die als Zielsequenz jeweils den Sequenz-Übergang aus einem Promotor (P-35S bzw. P-nos) in das nptII-Gen amplifizieren. Außerdem wird ein Gen-spezifisches real-time-PCR Verfahren für den Nachweis von DNA-Sequenzen des nos-Promotors aus Agrobacterium tumefaciens präsentiert.

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Copyright information

© Springer Basel AG 2010

Authors and Affiliations

  1. 1.Hessisches LandeslaborKasselGermany

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