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| Copy Number Estimation and Quantitative Analysis of Transgenic Rice Kefeng 6 Through Real-time PCR Strategies |
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Abstract Establishment of a sensitive and accurate quantitative labeling system for transgenic components of Genetically Modified Organisms (GMOs) is a crucial step to employ the biosafety management. Real-time quantitative PCR methods have been applied to quantitatively detect the transgenic components as a commonly used technique. By using this method, three inserted copies were detected in Kefeng-6, an insecticidal transgenic rice(Oryza sativa) harboring bivalent gene (cry1Ac+CpTi). Via using absolute quantitative and relative quantitative method to detect the inserted cry1Ac genes and its event-specific sequence in Kefeng 6, we found out that the detection both inserted genes and its event-specific sequence could meet the needs for the quantitative detection. As for 100 ng of genomic DNA, In relative quantitative PCR, the detection threshold of relative amount of cry1Ac gene and its specific flanking sequence was 0.1% and 0.5%, respectively; In absolute quantitative PCR, the detection threshold of the event-specific for Kefeng 6 was approximately five copies. We validated the accuracy of above methods by detecting four mixed samples with known GM contents. To quantitate the GM ratio of a product, the relative quantitation results were not affected either by multi-copy gene or single-copy gene. This study could provide a reliable reference to set up the threshold in quantitative detection of transgenic product.
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Received: 29 July 2011
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SHEN Xue-Jing, ZHANG Er-Qin, XU Qian-Ru, LIU Lin-Ke, WAN Bo, LIU Yun-Chao, SUN Ya-Ning, ZHAO Dong, NIU Xiang-Xiang, DENG Rui-Guang, ZHANG Gai-Ping. Expression of Rabies virus G Protein in Rice (Oryza sativa) and Identification of Its Genetic Stability[J]. 农业生物技术学报, 2019, 27(2): 204-211. |
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