转基因玉米2A-5特异性PCR方法的建立

根据转基因玉米2A-5的旁侧序列信息,设计并筛选出最佳特异性引物及Taqman探针组合2A-5-5-QF8、2A-5-5-QR8、2A-5-5-QP8,优化了该引物探针组合的反应体系,建立了转基因玉米2A-5转化体特异性PCR检测方法。将特异性引物及Taqman探针组合用于qRT-PCR和ddPCR技术,研究了转基因玉米2A-5转化体的定量检测方法,发现PCR定量检测转基因含量与测定样品转基因含量之间呈高度正相关。研究获得的转基因玉米2A-5转化体特异性PCR检测方法及定量qRT-PCR、ddPCR检测方法具有较高的特异性、准确性和灵敏度,是今后准确、高效检测2A-5及其产品的有效方法之一,同时也为我国转基因生物安全监管提供了良好的技术支撑。

Establishment of Event-specific PCR Detection Method of Transgenic Maize Line 2A-5

According to the flanking sequence information of transgenic maize 2A-5, the optimal specific primer and TaqMan probe combination 2A-5-5-QF8, 2A-5-5-QR8, 2A-5-5-QP8 were designed and screened. The reaction system of the combinations was optimized as well, and the specific PCR detection method of transgenic event 2A-5 was established. The quantitative detection method of transgenic maize 2A-5 was studied by qRT-PCR and ddPCR. It was found that the combination of specific primers and TaqMan probe could complete the quantitative detection well. There was a high positive correlation between the content of transgenic components obtained by PCR reaction and the content of the sample. The specific PCR detection method and quantitative qRT-PCR and ddPCR detection method of transgenic maize 2A-5 in this study can detect 2A-5 with high specificity, accuracy and sensitivity, which is one of the effective methods to detect 2A-5 and its products accurately and efficiently in the future, and it also can provide good technical support for the safety supervision of genetically modified organisms in China.