Key Laboratory of Oil Crop Biology of the Ministry of Agriculture, Oil Crops Research Institute, Chinese Academy of Agricultural Sciences , No. 2 Xudong 2nd Road, Wuhan 430062, People's Republic of China.
J Agric Food Chem. 2013 Jun 26;61(25):5953-60. doi: 10.1021/jf401339k. Epub 2013 Jun 14.
In this study, a collaborative trial of validating a real-time PCR method for the TT51-1 rice event was organized, including six participating laboratories. In this validation, serially diluted solutions from homogeneous genomic DNA of the TT51-1 event were used to construct standard curves of the TT51-1 event and phospholipase D (PLD) assays. The PCR efficiency was 95%, and the R(2) coefficient was 0.99 for the TT51-1 system. The mean quantitative values for blind samples containing 0.1%, 0.5% 1%, 5%, and 10% (w/w) TT51-1 corresponded to 0.1%, 0.51%, 1.06%, 4.83%, and 9.62%, respectively, with a bias (%) ranging from -3.77% to 5.87%. The repeatability and reproducibility were all below 25% across the entire dynamic range. Furthermore, the measurement uncertainties of the quantitative results were estimated to be 0.10%, 0.20%, 0.40%, 1.76%, and 3.52% (w/w) for the tested samples. Both the LOD and LOQ were calculated to be 0.22%. This collaborative trial demonstrated that the TT51-1 method produces reliable, comparable, and reproducible results for a given sample set and can be adopted as a detection standard for testing laboratories.
本研究组织了 TT51-1 水稻事件实时 PCR 方法的验证合作试验，共有 6 个参与实验室。在此次验证中，使用 TT51-1 事件同质基因组 DNA 的连续稀释溶液来构建 TT51-1 事件和磷脂酶 D（PLD）检测的标准曲线。TT51-1 体系的 PCR 效率为 95%，R(2)系数为 0.99。含有 0.1%、0.5%、1%、5%和 10%（w/w）TT51-1 的盲样定量值分别为 0.1%、0.51%、1.06%、4.83%和 9.62%，偏差（%）在-3.77%至 5.87%之间。在整个动态范围内，重复性和再现性均低于 25%。此外，对测试样品的定量结果的测量不确定度估计为 0.10%、0.20%、0.40%、1.76%和 3.52%（w/w）。LOD 和 LOQ 均计算为 0.22%。该合作试验表明，TT51-1 方法可为给定的样品集提供可靠、可比和可重复的结果，可作为检测实验室的检测标准。
