Event-specific Method for the Quantification of Soybean FG72 Using Real-time PCR: Validation Report and Validated Method

In line with its mandate the European Union Reference Laboratory for GM food and Feed (EU-RL GMFF), in collaboration with the European Network of GMO Laboratories (ENGL), has validated an event-specific PCR method for detecting and quantifying soybean event FG72 (unique identifier MST-FGØ72-2). The validation study was conducted according to the EU-RL GMFF validation procedure [http://gmo-crl.jrc.ec.europa.eu/guidancedocs.htm] and internationally accepted guidelines, involving 12 laboratories. In accordance with current EU legislation1, Bayer CropScience has provided the detection method and the samples (genomic DNA from soybean seeds harbouring the FG72 event as positive control DNA, genomic DNA from conventional soybean seeds as negative control DNA). The EU-RL GMFF prepared the validation samples (calibration samples and blind samples at test GM percentage [DNA/DNA]), organised an international collaborative study and analysed the results. The study confirms that the method meets the method performance requirements as established by the EU-RL and the ENGL detailing the provisions of Annex I-2.C.2 to Regulation (EC) No 641/2004 and it fulfils the analytical requirements of Regulation (EU) No 619/2011. This report is published at http://gmo-crl.jrc.ec.europa.eu/statusofdoss.htm.JRC.I.3-Molecular Biology and Genomic

Report on the Single-laboratory Validation of a PCR-based Detection Method for Identification of Florigene™ 26407 GM Carnation

Suntory Holdings Ltd has submitted an application for marketing (C/NL/09/02) of a genetically modified carnation line 26407 (Unique identifier: IFD-26407-2). In this context, the European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF) was asked to carry out a singlelaboratory validation of the performance of a polymerase chain reaction (PCR)-based method for detecting and identifying the carnation GM line 26407, developed by the applicant. This report describes the results of this validation, carried out by the EU-RL GMFF with control samples provided by the applicant. The method is a duplex end-point PCR, where a carnation (taxon) target and a transgenic sequence are detected simultaneously. The limit of detection (LOD) of the method has been established to be at least 10 copies for the taxon-specific target and between 50 and 10 copies for the GM target, based on haploid genome copy number. The event-specificity of the method was assessed by the applicant as being sufficient. The EU-RL could verify that the taxon-specific primers correctly detect the endogenous gene target in genomic DNA of a conventional carnation line (negative control) and in the genomic DNA of the GM carnation line, while the GM target is detected by the GM specific primers only in genomic DNA of 26407 GM line (positive control).JRC.I.3-Molecular Biology and Genomic

Report on the In-house Validation of a DNA Extraction Method from Soybean Grains and Validated Method

In accordance with relevant EU legislation , Dow AgroSciences LLC provided to the European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF) a DNA extraction method for soybean grains and the relevant samples (ground soybean grains). In line with its mandate , the EU RL GMFF has conducted an in-house validation of this DNA extraction method. To this end it tested the DNA extraction method on the samples provided and evaluated its performance in terms of DNA yield, integrity and quality. The in-house validation study confirmed that the method meets the method performance requirements as established by the EU-RL GMFF and the ENGL , and that it satisfies the provisions of Annex I-2.C.2 to Regulation (EC) No 641/2004. The method is therefore fit for the purpose of producing soybean DNA of suitable quantity and quality for subsequent PCR-based analysis.JRC.I.3-Molecular Biology and Genomic

Event-specific Method for the Quantification of Soybean DAS-68416-4 Using Real-time PCR: Validation report

In line with its mandate the European Union Reference Laboratory for GM Food and Feed (EU RL GMFF), in collaboration with the European Network of GMO Laboratories (ENGL), has validated an event-specific polymerase chain reaction (PCR) method for detecting and quantifying soybean event DAS-68416-4 (unique identifier DAS-68416-4). The validation study was conducted according to the EU-RL GMFF validation procedure (http://gmo-crl.jrc.ec.europa.eu/guidancedocs.htm) and internationally accepted guidelines. In accordance with current EU legislation , Dow AgroSciences LLC provided the detection method and the positive and negative control samples (genomic DNA extracted from soybean kernels harbouring the DAS-68416-4 event as positive control DNA, genomic DNA extracted from conventional soybean kernels as negative control DNA). The EU-RL GMFF prepared the validation samples (calibration samples and blind samples at different GM percentage [DNA/DNA]), organised an international collaborative study and analysed the results. The study confirms that the method meets the method performance requirements as established by the EU-RL GMFF and the ENGL and according to Annex I-2.C.2 to Regulation (EC) No 641/2004 and it fulfils the analytical requirements of Regulation (EU) No 619/2011JRC.I.3-Molecular Biology and Genomic

Report on the single-laboratory validation of a PCR-based Detection Method for Identification of Florigene™ IFD-25958-3 GM Carnation -Validation Report and Validated Method

In the context of the application for marketing submitted by Florigene Pty Ltd for a genetically modified carnation line (C/NL/09/01) IFD-25958-3, the European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF) has carried out a single-laboratory validation to assess the performance of a polymerase chain reaction (PCR)-based detection method for detecting and identifying the carnation GM line IFD-25958-3. This report describes the results of tests carried out by the EU-RL GMFF on control samples provided by the method developer and according to the detection method described by the applicant. The taxon-specific method correctly detects the endogenous gene target in genomic DNA of a conventional carnation line (negative control) and in the genomic DNA of the GM carnation line; the same method can also detect the GM target DNA in IFD-25958-3 GM line (positive control) in the experimental conditions described in this report. The Limit of Detection (LOD) of the method has been estimated to be at least 50 copies for the taxon-specific gene and at least 100 copies for the GM insert, based on haploid genome copy number.JRC.I.3-Molecular Biology and Genomic

Event-specific Method for the Quantification of Maize DAS-40278-9 by Real-time PCR - Validation Report and Validated Method

In line with its mandate1 the European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF), in collaboration with the European Network of GMO Laboratories (ENGL), has validated an event-specific polymerase chain reaction (PCR) method for detecting and quantifying maize event DAS-40278-9 (unique identifier DAS-4Ø278-9). The validation study was conducted according to the EU-RL GMFF validation procedure [http://gmo-crl.jrc.ec.europa.eu/guidancedocs.htm] and the internationally accepted guidelines2. In accordance with current EU legislation1, Dow AgroSciences LLC has provided the detection method and the positive and negative control samples (genomic DNA from maize seeds harbouring the DAS-40278-9 event as positive control DNA, genomic DNA from conventional maize seeds as negative control DNA). The EU-RL GMFF prepared the validation samples (calibration samples and blind samples at different GM percentage [DNA/DNA]), organised an international collaborative study and analysed the results. The study confirms that the method meets the method performance requirements as established by the EU-RL GMFF and the ENGL in Annex I-2.C.2 to Regulation (EC) No 641/20041 and it fulfils the analytical requirements of Regulation (EU) No 619/2011. This report is published at http://gmo-crl.jrc.ec.europa.eu/statusofdoss.htm.JRC.I.3-Molecular Biology and Genomic

Event-specific Method for the Quantification of Soybean CV127 using Real-time PCR

The European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF), established by Regulation (EC) No 1829/2003, has carried out a validation study to assess the performance of a quantitative event-specific method on the soybean event CV127 (unique identifier BPS-CV127-9). The collaborative trial was conducted according to internationally accepted guidelines. In accordance to Regulation (EC) No 1829/2003 of 22 September 2003 on genetically modified food and feed and to Regulation (EC) No 641/2004 of 6 April 2004 on detailed rules for the implementation of Regulation (EC) No 1829/2003, BASF Plant Science provided the detection method and the control samples. The EU-RL GMFF prepared the validation samples [calibration samples and blind samples at unknown GM percentages (DNA/DNA)].The results of the international collaborative trial met the European Network of GMO Laboratories (ENGL) method performance requirements (http://gmocrl.jrc.ec.europa.eu/guidancedocs.htm). The method is therefore considered applicable to the control samples provided, in accordance with the requirements of Annex I – 2.C.2 to Regulation (EC) No 641/2004.JRC.I.3-Molecular Biology and Genomic

Event-specific Method for the Quantification of Maize MON87460 Using Real-time PCR: Validation Report and Validated Method

The European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF), established by Regulation (EC) No 1829/2003, in collaboration with the European Network of GMO Laboratories (ENGL), has carried out a collaborative study to assess the performance of a quantitative event-specific method to detect and quantify the MON 87460 transformation event in maize DNA (unique identifier MON-8746Ø-4). The collaborative study was conducted according to internationally accepted guidelines (1, 2). In accordance with Regulation (EC) No 1829/2003 of 22 September 2003 on genetically modified food and feed and with Regulation (EC) No 641/2004 of 6 April 2004 on detailed rules for the implementation of Regulation (EC) No 1829/2003, Monsanto Company provided the detection method and the samples (genomic DNA extracted from homogenised seeds containing the transformation event and from conventional homogenised seeds). The EU-RL GMFF prepared the validation samples (calibration samples and blind samples at defined GM percentages [DNA/DNA], unknown to laboratories participating to the collaborative study). The collaborative trial involved twelve laboratories from ten European countries. The results of the international collaborative study met the ENGL performance requirements. The method is, therefore, considered applicable to the control samples provided, in accordance with the requirements of Annex I-2.C.2 to Commission Regulation (EC) No 641/2004. The results of the collaborative study are made publicly available at http://gmo-crl.jrc.ec.europa.eu/.JRC.I.3-Molecular Biology and Genomic

Event-specific Method for the Quantification of Maize 98140 by Real-time PCR

The European Union Reference Laboratory for GM Food and Feed (EU-RL GMFF), established by Regulation (EC) No 1829/2003, has carried out a validation study to assess the performance of a quantitative event-specific method on the maize event 98140 (unique identifier DP-098140-6). The collaborative trial was conducted according to internationally accepted guidelines. In accordance to Regulation (EC) No 1829/2003 of 22 September 2003 on genetically modified food and feed and to Regulation (EC) No 641/2004 of 6 April 2004 on detailed rules for the implementation of Regulation (EC) No 1829/2003, Pioneer Overseas Corporation provided the detection method and the control samples. The EU-RL GMFF prepared the validation samples [calibration samples and blind samples at unknown GM percentages(DNA/DNA)]. The results of the international collaborative trial met the European Network of GMO Laboratories (ENGL) method performance requirements (http://gmocrl.jrc.ec.europa.eu/guidancedocs.htm). The method is therefore considered applicable to the control samples provided, in accordance with the requirements of Annex I – 2.C.2 to Regulation (EC) No 641/2004.JRC.I.3-Molecular Biology and Genomic

Report on the Verification of the Performance of a MON810 Event-specific Method on Maize Line MON810 Using Real-time PCR

The JRC as Community Reference Laboratory for GM Food and Feed (CRL-GMFF) (see Regulation EC No 1829/2003), has carried out an in-house verification study to assess the performance of the MON810 method to detect and quantify the MON810 transformation event in maize DNA (unique identifier MON-¿¿810-6). The method has previously undergone a full validation on samples represented by certified reference material. The present verification was conducted in order to verify the performance of the validated method on the control samples provided by the applicant as requested by Annex I.2.C.2 to Regulation (EC) No 641/2004 stating that ¿The method shall be applicable to samples of the food or feed, to the control samples and to the reference material, which is referred to in Articles 5(3)(j) and 17(3)(j) of Regulation (EC) No 1829/2003.¿ The study was conducted according to internationally accepted guidelines (1,2). In accordance with Regulation (EC) No 1829/2003 of 22 September 2003 on genetically modified food and feed and to Regulation (EC) No 641/2004 of 6 April 2004 on detailed rules for the implementation of Regulation (EC) No 1829/2003, the CRL-GMFF carried out a verification of the event-specific detection method previously validated by the Federal Institute for Risk Assessment (BfR) in collaboration with the American Association of Cereal Chemists (AACC), Joint Research Centre (JRC) of the European Commission (EC), Institute for Reference Material and Measurement (IRMM), the Institute for Health and Consumer Protection (IHCP) and GeneScan, Berlin; Monsanto Company provided the control samples (MON810 maize seeds and conventional maize seeds) used in the verification. The JRC prepared the in-house verification samples (calibration samples and blind samples at different GM percentages). The results of the in-house verification study were evaluated with reference to ENGL method performance requirements (http://gmo-crl.jrc.ec.europa.eu/guidancedocs.htm) and to the results of the full validation (http://gmo-crl.jrc.ec.europa.eu/statusofdoss.htm). The results of CRL-GMFF in-house verification study are made publicly available at http://gmo-crl.jrc.ec.europa.eu/statusofdoss.htm).JRC.DDG.I.4-Molecular biology and genomic