Jacchia Sara, Nardini Elena, Bassani Niccolò, Savini Christian, Shim Jung-Hyun, Trijatmiko Kurniawan, Kreysa Joachim, Mazzara Marco
†Molecular Biology and Genomics Unit, Institute for Health and Consumer Protection, European Commission Joint Research Centre, Via E. Fermi 2749, 21027 Ispra, Varese, Italy.
‡Plant Breeding, Genetics and Biotechnology Division, International Rice Research Institute, Los Baños, 4031 Laguna, Philippines.
J Agric Food Chem. 2015 May 27;63(20):4954-65. doi: 10.1021/acs.jafc.5b00951. Epub 2015 May 13.
This article describes the international validation of the quantitative real-time polymerase chain reaction (PCR) detection method for Golden Rice 2. The method consists of a taxon-specific assay amplifying a fragment of rice Phospholipase D α2 gene, and an event-specific assay designed on the 3' junction between transgenic insert and plant DNA. We validated the two assays independently, with absolute quantification, and in combination, with relative quantification, on DNA samples prepared in haploid genome equivalents. We assessed trueness, precision, efficiency, and linearity of the two assays, and the results demonstrate that both the assays independently assessed and the entire method fulfill European and international requirements for methods for genetically modified organism (GMO) testing, within the dynamic range tested. The homogeneity of the results of the collaborative trial between Europe and Asia is a good indicator of the robustness of the method.
本文描述了用于检测黄金大米2号的定量实时聚合酶链反应（PCR）检测方法的国际验证情况。该方法包括一个扩增水稻磷脂酶Dα2基因片段的分类群特异性检测法，以及一个基于转基因插入片段与植物DNA之间3'连接处设计的事件特异性检测法。我们以单倍体基因组当量制备的DNA样本，对这两种检测法分别进行了绝对定量验证，以及将它们结合起来进行相对定量验证。我们评估了这两种检测法的准确性、精密度、效率和线性，结果表明，在测试的动态范围内，单独评估的这两种检测法以及整个方法均符合欧洲和国际对转基因生物（GMO）检测方法的要求。欧洲和亚洲之间合作试验结果的一致性是该方法稳健性的良好指标。
