Detection of Genetically Modified Canola Using Multiplex PCR Coupled with Oligonucleotide Microarray Hybridization

• Published in: Journal of agricultural and food chemistry 2008-08, Vol.56 (16), p.6791-6800
• Main Authors: Schmidt, Anna-mary, Sahota, Robert, Pope, Derek S, Lawrence, Tracy S, Belton, Mark P, Rott, Michael E
• Format: Article
• Language: English
• Subjects: Analytical Methods; Biological and medical sciences; Brassica napus; Brassica napus - classification; Brassica napus - genetics; Brassica napus var. napus; Canola; DNA fingerprinting; DNA probes; DNA, Plant - analysis; feed composition; feeds; Food industries; Fundamental and applied biological sciences. Psychology; General aspects; genetically modified; lines; Methods of analysis, processing and quality control, regulation, standards; microarray technology; multiplex PCR; nucleic acid hybridization; Oligonucleotide Array Sequence Analysis; oligonucleotide microarray; Oligonucleotide Probes; Plants, Genetically Modified - genetics; polymerase chain reaction; Polymerase Chain Reaction - methods; rapid methods; Sensitivity and Specificity; transgenes; transgenic plants
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/jf800137q

A rapid method was developed for concurrent screening of transgenic elements in GM canola. This method utilizes a single multiplex PCR coupled with an oligonucleotide DNA array capable of simultaneously detecting the 12 approved GM canola lines in Canada. The assay includes construct-specific elements for identification of approved lines, common elements (e.g., CaMV 35S promoter, Agrobacterium tumefaciens nos terminator, or nptII gene) for screening of approved or unapproved lines, a canola-specific endogenous gene, and endogenous genes from heterologous crops to serve as additional controls. Oligonucleotide probes were validated individually for functionality and specificity by amplification of specific transgene sequences from appropriate GM canola lines corresponding to each probe sequence, and hybridization of amplicons to the array. Each target sequence hybridized to its corresponding oligonucleotide probe and no significant cross-hybridization was observed. The limit of detection was examined for the GM lines GT73, T45, and MS8/RF3, and was determined to be 0.1%, 0.1%, and 0.5%, respectively, well within the European food and feed labeling threshold level of 0.9% for approved GM product. Practically, the method was demonstrated to be effective for the detection of GM canola in several types of animal feed, as well as in commercial canola meal.