title: Development and validation of event-specific quantitative PCR method for genetically modified maize MIR604.
abstract: A GM maize event, MIR604, has been widely distributed and an analytical method to quantify its content is required to monitor the validity of food labeling. Here we report a novel real-time PCR-based quantitation method for MIR604 maize. We developed real-time PCR assays specific for MIR604 using event-specific primers designed by the trait developer, and for maize endogenous starch synthase IIb gene (SSIIb). Then, we determined the conversion factor, which is required to calculate the weight-based GM maize content from the copy number ratio of MIR604-specific DNA to the endogenous reference DNA. Finally, to validate the developed method, an interlaboratory collaborative trial according to the internationally harmonized guidelines was performed with blind samples containing MIR604 at the mixing levels of 0, 0.5, 1.0, 5.0 and 10.0%. The reproducibility (RSDr) of the developed method was evaluated to be less than 25%. The limit of quantitation of the method was estimated to be 0.5% based on the ISO 24276 guideline. These results suggested that the developed method would be suitable for practical quantitative analyses of MIR604 maize.
authors.affiliation: National Food Research Institute, National Agriculture and Food Research Organization, Tsukuba, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Mano
authors.forename: Satoshi
authors.initials: S
authors.lastname: Furui
authors.forename: Kaori
authors.initials: K
authors.lastname: Takashima
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Koiwa
authors.forename: Satoshi
authors.initials: S
authors.lastname: Futo
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Minegishi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Akiyama
authors.forename: Reiko
authors.initials: R
authors.lastname: Teshima
authors.forename: Takeyo
authors.initials: T
authors.lastname: Kurashima
authors.forename: Reona
authors.initials: R
authors.lastname: Takabatake
authors.forename: Kazumi
authors.initials: K
authors.lastname: Kitta
mesh_terms: D030841:Food, Genetically Modified; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D003313:Zea mays / Q000235:genetics