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为了方便快速地检测转cry1Ac基因抗虫水稻的转基因成分,本研究以转cry1Ac基因水稻‘华恢1号’为主要研究对象,根据环介导等温扩增技术的原理,针对人工改造的cry1Ac晶体蛋白毒素基因的6个区段设计4条特异性引物,在65℃条件下,利用链置换DNA聚合酶等温扩增1h,分别通过荧光显色和凝胶电泳完成对转基因作物的检测工作,同时对2种转基因作物和5种非转基因作物进行LAMP检测以检验该方法的特异性。结果表明,该LAMP方法能有效地检测含有cry1Ac的转基因作物,检测结果与常规PCR方法结果一致,灵敏度是PCR方法的10倍,达到0.01%,并且具有高特异性,表明该检测方法适合转cry1Ac基因抗虫作物的快速检测。
In order to detect the transgenic components of insect-resistant rice transformed with cry1Ac gene conveniently and rapidly, we studied the cry1Ac transgenic rice “Huahui No.1” as the main research object. According to the principle of ring-mediated isothermal amplification, Four specific primers were designed according to six segments of the protein toxin gene. The PCR products were isothermally amplified by strand displacement DNA polymerase at 65 ℃ for 1 h. The detection of transgenic crops was completed by fluorescence and gel electrophoresis. LAMP assays were performed on 2 transgenic and 5 non-transgenic crops to test the specificity of the method. The results showed that the LAMP method can effectively detect cry1Ac-containing transgenic crops. The results of the LAMP assay are consistent with those of the conventional PCR method. The sensitivity of the LAMP method is ten times that of the PCR method with a specificity of 0.01% and high specificity, indicating that the detection method is suitable for cry1Ac Rapid Detection of Gene-resistant Crops.