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首页> 外文会议>Conference on Nanoscale Imaging, Sensing, and Actuation for Biomedical Applications; 20080121-23; San Jose,CA(US) >Quantitative detection of antibiotic resistance genes using magnetic/luminescent core-shell nanoparticles
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Quantitative detection of antibiotic resistance genes using magnetic/luminescent core-shell nanoparticles

机译：使用磁性/发光核-壳纳米粒子定量检测抗生素抗性基因

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摘要

Nanoscale magnetic/luminescent core-shell particles were used for DNA quantification in a hybridization-in-solution format. We demonstrated a simple, high-throughput, and non-PCR based DNA assay for quantifying antibiotic resistance gene tetQ. Fe_3O_4/Eu:Gd_2O_3 nanoparticles (NPs) synthesized by spray pyrolysis were biofunctionalized by passive adsorption of NeutrAvidin. Following immobilization of biotinylated probe DNA on the particles' surfaces, target dsDNA and signaling probe DNA labeled with Cy3 were hybridized with NPs-probe DNA. Hybridized DNA complexes were separated from solution by a magnet, while non-hybridized DNA remained in solution. A linear quantification (R~2 = 0.99) of a target tetQ gene was achieved based on the normalized fluorescence (Cy3/NPs) of DNA-NP hybrids. A real-time qPCR assay was used for evaluation of the NPs assay sensitivity and range of quantification. The quantity of antibiotic resistance tetQ genes in activated sludge microcosms, with and without addition of tetracycline or triclosan has been determined, indicating the potential of the optimized assay for monitoring the level of antibiotic resistance in environmental samples. In addition, the tetQ gene copy numbers in microcosms determined by NP-hybridization were well correlated with the numbers measured by real-time qPCR assay (R~2 = 0.92).
机译：纳米级磁性/发光核-壳颗粒用于溶液杂交形式的DNA定量。我们展示了一种简单，高通量且基于非PCR的DNA分析方法，用于定量抗生素抗性基因tetQ。喷雾热解合成的Fe_3O_4 / Eu：Gd_2O_3纳米颗粒（NPs）通过NeutrAvidin的被动吸附进行生物功能化。将生物素化的探针DNA固定在颗粒表面后，将用cy3标记的目标dsDNA和信号探针DNA与NPs-probe DNA杂交。用磁铁将杂交的DNA复合物从溶液中分离出来，而未杂交的DNA保留在溶液中。基于DNA-NP杂种的归一化荧光（Cy3 / NPs），实现了目标tetQ基因的线性定量（R〜2 = 0.99）。实时定量PCR检测用于评估NP检测灵敏度和定量范围。已确定在有或没有添加四环素或三氯生的情况下，活性污泥微观世界中的抗生素抗性tetQ基因的数量，表明优化的检测方法可能用于监测环境样品中的抗生素抗性水平。此外，通过NP杂交确定的微观世界中的tetQ基因拷贝数与通过实时qPCR测定法测得的数目有很好的相关性（R〜2 = 0.92）。

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