Development and in-house validation of the event-specific qualitative and quantitative PCR detection methods for genetically modified cotton MON15985

• Published in: Journal of the science of food and agriculture Vol. 90; no. 3; pp. 402 - 408
• Main Authors: Jiang, Lingxi, Yang, Litao, Rao, Jun, Guo, Jinchao, Wang, Shu, Liu, Jia, Lee, Seonghun, Zhang, Dabing
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 01.02.2010; Wiley; John Wiley and Sons, Limited
• Subjects: Base Sequence; Biological and medical sciences; Cotton; Deoxyribonucleic acid; detection; DNA; DNA Primers; DNA probes; DNA, Plant - analysis; event-specific; Food industries; Food science; Fundamental and applied biological sciences. Psychology; General aspects; genetically modified cotton; Genome, Plant; Genomics; Gossypium - genetics; Gossypium hirsutum; Haploidy; in-house validation; Limit of Detection; Methods of analysis, processing and quality control, regulation, standards; MON15985; Plants, Genetically Modified - genetics; polymerase chain reaction; Polymerase Chain Reaction - methods; qualitative analysis; quantitative analysis; Studies; transgenic plants; Polymerase chain reaction; Validation; Analysis method; Development; Control method; MON15985; event-specific; genetically modified cotton; Molecular biology; Detection; Genetically modified organism; in-house validation
• ISSN: 0022-5142; 1097-0010
• DOI: 10.1002/jsfa.3829

BACKGROUND: To implement genetically modified organism (GMO) labeling regulations, an event-specific analysis method based on the junction sequence between exogenous integration and host genomic DNA has become the preferential approach for GMO identification and quantification.RESULTS: In this study, specific primers and TaqMan probes based on the revealed 5′-end junction sequence of GM cotton MON15985 were designed, and qualitative and quantitative polymerase chain reaction (PCR) assays were established employing the designed primers and probes. In the qualitative PCR assay, the limit of detection (LOD) was 0.5 g kg⁻¹ in 100 ng total cotton genomic DNA, corresponding to about 17 copies of haploid cotton genomic DNA, and the LOD and limit of quantification (LOQ) for quantitative PCR assay were 10 and 17 copies of haploid cotton genomic DNA, respectively. Furthermore, the developed quantitative PCR assays were validated in-house by five different researchers. Also, five practical samples with known GM contents were quantified using the developed PCR assay in in-house validation, and the bias between the true and quantification values ranged from 2.06% to 12.59%.CONCLUSION: This study shows that the developed qualitative and quantitative PCR methods are applicable for the identification and quantification of GM cotton MON15985 and its derivates.