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[Establishment of a field visual detection method for genetically modified maize 'Shuangkang'12-5 by fluorescence
Yu Chen1, Xiao-Yun Chen1, Cheng Peng1
1tate Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-products, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China.
Abstract:
The genetically modified (GM) maize 'Shuangkang'12-5 has good insect resistance and herbicide tolerance, which is one of the first series of GM maizes obtained a safety certificate in China, and it has broad application prospect in the future. This study established an on-site rapid detection method for GM 'Shuangkang'12-5 based on recombinase polymerase amplification (RPA) technology, which primes and probe were designed according to the specific flank sequence. Then the best combination of primers and probe was obtained through a screeing process. The amplification results of fluorescence RPA can be directly visualized under blue light. The results showed that the visual detection system of GM 'Shuangkang'12-5 with high specificity, and the detection sensitivity of the method could reached 10 copies. Further research found that the RPA amplification system had a wide range of temperature (34℃-46℃). According to this property, the common self-heating warm pastes on the market were used replace the traditional heating instruments to stimulate the RPA.The results showed that the self-heating warm paste meets the temperature requirement of the RPA system. Finally, we combined the self-heating warm pastes with the RPA visual detection system to conduct on-site detection of GM 'Shuangkang'12-5, and compared the results with the detection results of qPCR. The detection showed that the results of on-site visual detection method established in this study were consistent with the detection results of the qPCR. Moreover, the visual detection method was more shorter in time and the final detection result was clear and easy to distinguish. The rapid on-site visual detection method for GM 'Shuangkang' 12-5 established in this study has high specificity, high sensitivity and convenience. It not only meets the needs of on-site rapid detection of GM 'Shuangkang'12-5, but also provides highlight for the development of other on-site rapid detection methods.
