title: Effect of Source of DNA on the Quantitative Analysis of Genetically Engineered Traits Using Digital PCR and Real-Time PCR.
abstract: Seven commercially available DNA extraction kits were compared with a cetyltrimethylammonium bromide (CTAB) method to determine the suitability of the extracted DNA for RainDrop digital PCR (dPCR) and real-time PCR (RT-PCR) quantification of OXY235 canola, FP967 flax, and DP305423 soybean (spiked at the 0.1% level). For the kits, the highest amount of DNA extracted from a 0.2 g sample was obtained using OmniPrep for Plant for flax and DNeasy mericon Food for canola and soybean. For canola, DNA extracted with the Fast ID Genomic DNA Extraction Kit, FastDNA Spin Kit, GM Quicker 2, NucleoSpin Food, and DNeasy mericon Food was suitable for dPCR and RT-PCR. For flax, DNA extracted with Fast ID, FastDNA Spin Kit, OmniPrep for Plant, and NucleoSpin Food was suitable for RT-PCR. However, only Fast ID yielded DNA suitable for dPCR. For soybean, DNA extracted with five and six of the seven DNA extraction kits was suitable for dPCR and RT-PCR, respectively. Overall, Fast ID provided reliable results regardless of species or analysis method used. Canola, flax, and soybean DNA extracted with the CTAB method and then purified were suitable for both dPCR and RT-PCR. This is the first report showing the effect of different DNA extraction methods on the absolute quantification of genetically engineered traits using dPCR.
authors.affiliation: Canadian Grain Commission, Grain Research Laboratory, Winnipeg, MB, Canada.
authors.forename: Tigst
authors.initials: T
authors.lastname: Demeke
authors.affiliation: Agriculture and Agri-Food Canada, Morden, MB, Canada.
authors.forename: Jemima
authors.initials: J
authors.lastname: Malabanan
authors.affiliation: Canadian Grain Commission, Grain Research Laboratory, Winnipeg, MB, Canada.
authors.forename: Michelle
authors.initials: M
authors.lastname: Holigroski
authors.affiliation: Canadian Grain Commission, Grain Research Laboratory, Winnipeg, MB, Canada.
authors.forename: Monika
authors.initials: M
authors.lastname: Eng
mesh_terms: D029741:Brassica rapa / Q000235:genetics; D018744:DNA, Plant / Q000032:analysis* / Q000302:isolation & purification; D002523:Edible Grain / Q000235:genetics; D019597:Flax / Q000235:genetics; D030821:Plants, Genetically Modified; D060888:Real-Time Polymerase Chain Reaction; D013025:Glycine max / Q000235:genetics
chemical_list: D018744:DNA, Plant