Qualitative and Quantitative Event-Specific PCR Detection Methods for Oxy-235 Canola Based on the 3′ Integration Flanking Sequence

• Published in: Journal of agricultural and food chemistry Vol. 56; no. 6; pp. 1804 - 1809
• Main Authors: Yang, Litao, Guo, Jinchao, Zhang, Haibo, Liu, Jia, Zhang, Dabing
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 26.03.2008
• Subjects: Analytical Methods; Base Sequence; Biological and medical sciences; Brassica napus var. napus; Brassica rapa - classification; Brassica rapa - genetics; canola; Cloning, Molecular; detection; detection limit; DNA, Plant - analysis; DNA, Plant - chemistry; event-specific; Fish and seafood industries; Food industries; Fundamental and applied biological sciences. Psychology; General aspects; genetically modified foods; Genetically modified organisms; Methods of analysis, processing and quality control, regulation, standards; molecular sequence data; new methods; nucleotide sequences; Oxy-235 canola; Plants, Genetically Modified - classification; Plants, Genetically Modified - genetics; polymerase chain reaction; Polymerase Chain Reaction - methods; quantitation limit; quantitative analysis; real-time PCR; recombinant DNA; Reproducibility of Results; Sensitivity and Specificity; TAIL-PCR; transgenic plants; event-specific; real-time PCR; Oxy-235 canola; TAIL-PCR; Genetically modified organisms; Bass; Edible fish; Real time; Polymerase chain reaction; Analysis method; Control method; Molecular biology; Detection; Genetically modified organism
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/jf073465i

As more genetically modified plant events are approved for commercialization worldwide, the event-specific PCR method has become the key method for genetically modified organism (GMO) identification and quantification. This study reveals the 3′ flanking sequence of the exogenous integration of Oxy-235 canola employing thermal asymmetric interlaced PCR (TAIL-PCR). On the basis of the revealed 3′ flanking sequence, PCR primers and TaqMan probe were designed and qualitative and quantitative PCR assays were established for Oxy-235 canola. The specificity and limits of detection (LOD) and quantification (LOQ) of these two PCR assays were validated to as low as 0.1% for the relative LOD of qualitative PCR assay; the absolute LOD and LOQ were low to 10 and 20 copies of canola genomic DNA in quantitative PCR assay, respectively. Furthermore, ideal quantified results were obtained in the practical canola sample detection. All of the results indicate that the developed qualitative and quantitative PCR methods based on the revealed 3′ integration flanking sequence are suitable for GM canola Oxy-235 identification and quantification.