Comparison of plasmid and genomic DNA calibrants for the quantification of genetically modified ingredients
Comparison of plasmid and genomic DNA calibrants for the quantification of genetically modified ingredients
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DOI：
10.1007/s00217-006-0376-z
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发表时间：
2006-12-01
影响因子：
3.3
通讯作者：
Harris, Neil
中科院分区：
文献类型：
--
作者：
Burns, Malcolm;Corbisier, Philippe;Harris, Neil
实时PCR是定量食品中转基因（GM）含量的首选方法。欧盟委员会最近的建议主张将转基因含量表示为每个目标分类单元特定序列的转基因目标DNA序列数量的百分比。因此，需要提供经拷贝数含量认证的参考材料，例如质粒形式的参考材料，以满足欧盟的这一建议。进行了一项实验室间试验，使用Roundup Ready（TM）大豆模型系统，比较质粒DNA与更传统的基因组DNA定量PCR校准方法的拷贝数。在三个国际实验室的重现性条件下，分析了48个随机实时定量PCR平板的数据。结果表明，在实验室试验中使用的特定质粒DNA提供了一个合适的替代基因组DNA用作校准品在GM定量。在当前研究中，质粒校准品在精密度和接近预期值方面的性能特征与其基因组等同物相同或更好。
Real-time PCR is the method of choice for the quantification of the genetically modified (GM) content of food. Recent EU Commission recommendations advocate expressing the GM content as the percentage of the number of GM target DNA sequences per target taxon specific sequence. The provision of reference materials certified for their copy number content, for example in the form of plasmids, are hence desirable to fulfil this EU recommendation. An inter-laboratory trial was conducted to compare plasmid DNA to the more traditional genomic DNA approach for quantitative PCR calibration in terms of copy numbers, using the model system of Roundup Ready (TM) soya. Data was analysed from 48 randomised real-time quantitative PCR plates under conditions of reproducibility across three international laboratories. Results demonstrated that the specific plasmid DNA used in the laboratory trial provided a suitable alternative to genomic DNA for use as a calibrant in GM quantification. In the current investigation, plasmid calibrants gave equal or better performance characteristics in terms of precision and closeness to the expected value, than their genomic equivalents.