Report on the Verification of the Performance of a Method for the Detection of Event MON71800 in Wheat Using Real-Time PCR

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Report on the Verification of the Performance of a Method for the Detection of Event MON71800 in Wheat Using Real-Time PCR
Abstract: 

Following the United States Department of Agriculture's (USDA) Animal and Plant Health Inspection Service (APHIS) announcement that test results confirmed the finding of unauthorised GM glyphosate-resistant wheat "volunteer" plants harbouring the event MON71800 on a farm in Oregon, the European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF) was requested to provide as soon as possible a method to test wheat consignments for the presence of this genetically modified organism (GMO) to the National Reference Laboratories (NRLs) for GMOs of the EU Member States. In response, the EU-RL put together a testing strategy, based on readily available screening tests which was published here (http://gmo-crl.jrc.ec.europa.eu/GM_wheat.htm). Upon request, Monsanto provided in May 2013 the EU-RL with the procedure ?Roundup Ready? Wheat MON71800 Event Specific Endpoint TaqMan? PCR with acc Internal Control for Seed Pools of 1:15? that had previously been made available to, and was used by USDA. The EU-RL GMFF tested this protocol on positive control samples consisting of MON71800 crude lysate, also provided by Monsanto. Our results can be summarised as follow: The method is apparently event-specific. Our specificity-tests did not show cross-reactivity on genomic DNA from a wide selection of similar GMO. The sensitivity of the method was found to be in agreement with previous findings of USDA, i.e. the relative limit of detection lies at 0.5% in a background of 301 ng of total wheat genomic DNA. The absolute limit of detection (LODabs) was determined between 5 and 10 copies of MON71800 target. The latter was not indicated by the USDA. For seed/grains the application of a sub-sampling strategy could allow detection below 0.5% but would require significant additional efforts, including the analysis of numerous sub-samples. Our tests also indicated that the duplex PCR system at the tested stage of optimisation is characterised by poor efficiency at increasing background DNA concentration in reaction. Based on the scientific evidence described in the present report, the EU-RL suggest that its testing strategy (http://gmo-crl.jrc.ec.europa.eu/GM_wheat.htm), making use of validated element and construct-specific methods and found to be more sensitive, is used to test for presence of MON71800 GM-wheat. The verified event specific method of Monsanto could be used to confirm positive findings at GM-target concentration equal or above 0.5% or it could be used for detection of GM-event MON71800 below 0.5% but it would require a costly sub-sampling strategy, which, in addition, is only possible in seeds/grains.

Authors
Authors: 
PATAK DENNSTEDT Alexandre, MAZZARA Marco, SAVINI Cristian, PETRILLO Mauro, ANGERS Alexandre, KREYSA Joachim, BOGNI Alessia
Publication Year
Publication Year: 
2013
Type

Type:

Appears in Collections
Appears in Collections: 
Institute for Health and Consumer Protection
Science Areas
JRC Institutes
Publisher
Publisher: 
Publications Office of the European Union
ISBN
ISBN: 
978-92-79-34671-2
ISSN
ISSN: 
1831-9424
DOI
URI
URI : 
Other Identifiers
Other Identifiers: 
EUR 26335
OPOCE LB-NA-26335-EN-N