Background To provide the consumer with choices of genetically modified organisms (GMO) or non-GMO, official food labeling systems were established in many countries. Because the threshold GMO content values were set to distinguish between “non-GMO” and “GMO” designations, GMO content quantification methods are required for ensuring the appropriateness of labeling. Objective As the number of GMOs is continuously increasing around the world, we set out to develop a low-cost, simple and less biased analytical strategy to cover all necessary detection targets. Methods Digital PCR methods are advantageous compared to the conventional quantitative real-time PCR methods. We developed a digital PCR-based GMO quantification method to evaluate the GMO content in maize grains. To minimize the analytical workload, we adopted multiplex digital PCR targeting the 35S promoter and the nopaline synthase terminator, which are genetic elements commonly introduced in many GMOs. Results Our method is significantly simpler and more precise than the conventional real-time PCR-based methods. Additionally, we found that this method enables quantification of the copy number of GMO DNA without double counting multiple elements (35S promoter and nopaline synthase terminator) tandemly placed in a recombinant DNA construct. Conclusion This is the first report on the development of a genetically modified maize quantification method using a multiplexed genetic element-specific digital PCR method. The tandem effect we report here is quite useful for reducing the bias in the analytical results. Highlights Multiplexed genetic element-specific digital PCR can simplify weight-based GMO quantification and thus should prove useful in light of the continuous increase in the number of GM events.

中文翻译：

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通过多重遗传元素特异性数字 PCR 进行简单、精确且偏差较小的 GMO 定量

背景 为了向消费者提供转基因生物（GMO）或非转基因生物的选择，许多国家建立了官方食品标签系统。由于设定了转基因含量阈值以区分“非转基因”和“转基因”标识，因此需要采用转基因含量量化方法来确保标签的适当性。目标随着全球转基因生物数量的不断增加，我们着手开发一种低成本、简单且偏差较小的分析策略，以涵盖所有必要的检测目标。方法 与传统的定量实时 PCR 方法相比，数字 PCR 方法具有优势。我们开发了一种基于数字 PCR 的 GMO 量化方法来评估玉米粒中的 GMO 含量。为了尽量减少分析工作量，我们采用了针对 35S 启动子和胭脂碱合酶终止子的多重数字 PCR，它们是许多转基因生物中常见的遗传元件。结果 我们的方法比传统的基于实时 PCR 的方法更简单、更精确。此外，我们发现这种方法能够量化 GMO DNA 的拷贝数，而无需重复计算串联放置在重组 DNA 构建体中的多个元素（35S 启动子和胭脂碱合酶终止子）。结论 这是关于使用多重遗传元件特异性数字 PCR 方法开发转基因玉米定量方法的第一份报告。我们在此报告的串联效应对于减少分析结果的偏差非常有用。