Development of taxon-specific sequences of common wheat for the detection of genetically modified wheat
Development of taxon-specific sequences of common wheat for the detection of genetically modified wheat
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DOI：
10.1021/jf0505731
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发表时间：
2005-08-10
影响因子：
6.1
通讯作者：
Hino, A
中科院分区：
文献类型：
--
作者：
Iida, M;Yamashiro, S;Hino, A
介绍了用于小麦DNA特异性检测和定量的定性和定量聚合酶链反应（PCR）系统。许多国家已颁布法规对含有转基因生物（GMO）的食品进行标识。PCR技术被广泛认为是转基因生物定性和定量检测的可靠和有用的技术。目的基因的扩增需要检测方法，扩增结果应与相应分类单元特异性参考基因的扩增结果进行比较，以获得可靠的结果。本文报道了普通小麦（Triticum aestivum L.）设计waxy-D1基因的特异性引物和TaqMan探针进行PCR分析。引物扩增出102 bp的产物（Wx 012）。结果表明，Wx 012基因序列是小麦的特异性序列，在19个小麦品种中，其定性PCR结果具有均一性，定量准确性沿着。在Southern印迹和实时PCR分析中，该序列显示单个或低拷贝数的基因。此外，利用wx 012引物和wx 012-T探针对小麦基因组进行定性和定量PCR，其检测限约为15个拷贝，且重复性可靠。因此，使用wx 012引物和wx 012-T探针的PCR系统被认为适合用作DNA分析（包括GMO测试）中的常见小麦特异分类群特异参考基因。
Qualitative and quantitative Polymerase Chain Reaction (PCR) systems aimed at the specific detection and quantification of common wheat DNA are described, Many countries have issued regulations to label foods that include genetically modified organisms (GMOs). PCR technology is widely recognized as a reliable and useful technique for the qualitative and quantitative detection of GMOs. Detection methods are needed to amplify a target GM gene, and the amplified results should be compared with those of the corresponding taxon-specific reference gene to obtain reliable results. This paper describes the development of a specific DNA sequence in the waxy-D1 gene for common wheat (Triticum aestivum L.) and the design of a specific primer pair and TaqMan probe on the waxy-D1 gene for PCR analysis. The primers amplified a product (Wx012) of 102 bp. It is indicated that the Wx012 DNA sequence is specific to common wheat, showing homogeneity in qualitative PCR results and very similar quantification accuracy along 19 distantly related common wheat varieties. In Southern blot and real-time PCR analyses, this sequence showed either a single or a low number of copy genes. In addition, by qualitative and quantitative PCR using wx012 primers and a wx012-T probe, the limits of detection of the common wheat genome were found to be about 15 copies, and the reproducibility was reliable. In consequence, the PCR system using wx012 primers and wx012-T probe is considered to be suitable for use as a common wheat-specific taxon-specific reference gene in DNA analyses, including GMO tests.