Collaborative trial validation of a construct-specific real-time PCR method for detection of genetically modified linseed event ‘CDC Triffid’ FP967

Abstract

A real-time PCR-based method for construct-specific detection of the genetically modified (GM) linseed event ’CDC Triffid’ FP967 originating from Canada has been validated in a collaborative trial. The construct-specific method amplifies a 105 bp long fragment of the transgenic insertion present in FP967 spanning the junction of the terminator region of the nopalin synthase gene from Agrobacterium tumefaciens (Tnos) to a sequence region coding for the dehydrofolate reductase gene (dfr) from a class I integron from Escherichia coli. This region is characteristic for the construct used to develop FP967. A total of 11 laboratories participated in the collaborative study. For PCR analysis, each laboratory received 14 DNA samples comprising 7 double-blind DNA samples. The samples consisted of two low GM-levels of FP967 DNA (10 or 50 copies per PCR), of DNA from two different GM-positive linseed products and of DNA from GM-negative linseed, potato and rapeseed materials, respectively. All but one of the FP967-positive DNA samples were detected correctly. No false-positive results were reported. The results demonstrate that the linseed event FP967 is detectable even at low copy number concentrations. The limit of detection (LOD) determined with plasmid DNA was shown to be at 5 copies of the Tnos–dfr sequence. The data provided show that the method can be applied successfully in different laboratories and is fit-for-purpose to test for the presence of the EU-unauthorised linseed event ‘CDC Triffid’ FP967.