转基因玉米实验室PCR检测体系的优化及应用

Establishment and Application of PCR Detection System of Genetically Modified Maize in Laboratory

为建立稳定的转基因玉米PCR检测体系,本实验采用正交设计法优化了玉米PCR反应体系,并对当地主要种植的16个玉米品种进行了转基因检测。结果显示Mg2+浓度对PCR扩增结果有较大影响,浓度低时影响扩增条带的特异性。降低d NTPs和引物浓度有助于扩增出较清晰的带型。3对定性PCR引物均可扩增出2个转基因玉米阳性样品的z SSIIb、Ca MV35S、NOS基因,而随机抽样的16个玉米样品均未检测到转基因目的片段。优化的PCR方法可作为转基因玉米实验室定性筛查的推荐方法,转基因玉米在本地的种植并不普遍。

Orthogonal design was used to establish the optimized PCR detection system of genetically modified maize in laboratory, with which 16 local maize varieties planted widely were detected. Results showed that Mg2+concentration had great influences on PCR amplification result, low concentration led to bands waning of non-specific amplification. Lower concentration of d NTPs and primers were optimized to amplify distinct bands.With three pairs of qualitative PCR primers, z SSIIb, Ca MV35 S and NOS were all amplified in two transgenic maize positive samples, whereas, in 16 randomly selected maize samples, all transgenic target fragments were not amplified. Thus, optimized PCR system could be used to screen genetically modified maize in laboratory, and transgenic maize variety are not widely planted locally.