Nanoparticle-based DNA biosensor for visual detection of genetically modified organisms
Nanoparticle-based DNA biosensor for visual detection of genetically modified organisms
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DOI：
10.1016/j.bios.2005.04.016
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发表时间：
2006-01-15
影响因子：
12.6
通讯作者：
Ioannou, PC
中科院分区：
文献类型：
--
作者：
Kalogianni, DP;Koraki, T;Ioannou, PC
尽管可以通过检测外源DNA或新蛋白质来完成对转基因生物（GMO）的原料和食品的筛选，但DNA是优选的分析物，因为其在食品加工期间具有上级稳定性。由于对快速可靠的转基因生物检测方法的需求日益增长，DNA生物传感器的开发变得越来越重要。我们报告了第一个DNA生物传感器在干试剂试纸配置的视觉检测和确认转基因相关序列的杂交在几分钟内。传感器是一次性的，不需要特殊的仪器。它检测存在于大多数转基因植物中的35S启动子和胭脂碱合酶（NOS）终止子序列。通过聚合酶链反应（PCR）扩增靶序列，并与带有寡聚（dA）尾的探针杂交（7分钟）。将生物素化产物应用于传感器，然后浸入适当的缓冲液中。缓冲液的迁移使与寡聚体（dT）缀合的金纳米颗粒再水化，所述寡聚体（dT）与寡聚体（dA）尾部杂交。在传感器的测试区，杂交体被固定化的链霉亲和素捕获，由于纳米颗粒的积累而产生特征红线。过量的纳米颗粒缀合物在对照区被固定化的寡（dA）链捕获。扩增的35S或NOS DNA在0.16 nM下可检测到。含有0.1%转基因成分的大豆粉标准物质在35和40个扩增循环后，35S和NOS序列分别清晰地检测到。该传感器还应用于来自各种来源的真实的样本。(c)2005 Elsevier B.V.保留所有权利。
Although screening of raw ingredients and food products for genetically modified organisms (GMO) may be accomplished by detecting either the exogenous DNA or the novel protein, DNA is the preferred analyte because of its superior stability during food processing. The development of DNA biosensors is of increasing importance due to the growing demand for rapid and reliable methods for GMO detection. We report the first DNA biosensor in a dry-reagent dipstick configuration for visual detection and confirmation of GMO-related sequences by hybridization within minutes. The sensor is disposable and does not require special instrumentation. It detects the 35S promoter and nopaline synthase (NOS) terminator sequences that are present in the majority of transgenic plants. The target sequences are amplified by the polymerase chain reaction (PCR) and hybridized (7 min) with probes bearing oligo(dA) tail. The biotinylated product is applied to the sensor followed by immersion in the appropriate buffer. Migration of the buffer rehydrates gold nanoparticles conjugated to oligo(dT), which hybridize with the oligo(dA) tails. The hybrids are captured by immobilized streptavidin at the test zone of the sensor giving a characteristic red line due to the accumulation of the nanoparticles. The excess of nanoparticle conjugates are captured at the control zone by immobilized oligo(dA) strands. Amplified 35S or NOS DNA is detectable at 0.16 nM. Soybean powder certified reference material with 0.1% GMO content is clearly detectable after 35 and 40 amplification cycles for 35S and NOS sequence, respectively. The sensor was also applied to real samples from various sources. (c) 2005 Elsevier B.V. All rights reserved.