关键词: 转基因玉米, 品系VCO-01981-5, 微滴式数字PCR, 定量检测

Abstract: In this paper, a duplex droplet digital polymerase chain reaction (ddPCR) method was developed to detect the content of unauthorized genetically modified (GM) maize event VCO-01981-5 based on a QX100 ddPCR platform. Towards this goal, PCR primers and TaqMan probes were designed based on the maize endogenous gene hmg and VCO-01981-5 event specific boundary sequences, and the probes were labeled with different chromophores so that the two targets could be detected in one single droplet. Analysis of the specificity of the method showed that both endogenous gene hmg and foreign sequence could be detected only in event VCO-01981-5. Moreover, the sensitivity, linearity and accuracy were evaluated and it was found that when relative standard deviation (RSD) was 25% or less, 5 copies of boundary sequences and 4 copies of gene hmg could be quantitatively detected, and the quantitative results (target copy) were highly correlated with the amount of DNA template up to 50 ng (R2 ≥ 0.99), with an average error of less than 10%. All the above results indicated that the established method in this study was very specific, stable, accurate and sensitive, and had a wide quantitative range in quantitative analysis of GM maize VCO-01981-5. Therefore, this method can be applied practically in quantitatively determining this GM maize event in imported and exported agricultural product and foods. Besides, this reported method can be used as a reference to establish a quantitative method for the detection of other GM maize events and GM events of other plants.

Key words: transgenic maize, event VCO-01981-5, droplet digital PCR, quantitative detection