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A Microfluidic Quantitative Polymerase Chain Reaction Method for the Simultaneous Analysis of Dozens of Antibiotic Resistance and Heavy Metal Resistance Genes
Environmental Science & Technology Letters ( IF 8.9 ) Pub Date : 2017-12-20 00:00:00 , DOI: 10.1021/acs.estlett.7b00552 Kyle D. Sandberg 1 , Satoshi Ishii 2, 3 , Timothy M. LaPara 1, 2
Environmental Science & Technology Letters ( IF 8.9 ) Pub Date : 2017-12-20 00:00:00 , DOI: 10.1021/acs.estlett.7b00552 Kyle D. Sandberg 1 , Satoshi Ishii 2, 3 , Timothy M. LaPara 1, 2
Affiliation
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This study developed, optimized, and demonstrated a microfluidic quantitative polymerase chain reaction (MF-qPCR) method for the simultaneous quantification of 39 antibiotic resistance genes (ARGs), five heavy metal resistance genes, three genes encoding the integrase of three different classes of integrons, and 16S rRNA genes (used as a measure of total bacterial biomass). Because the volume of the template is much smaller with MF-qPCR (a few nanoliters) than with conventional qPCR, a preamplification step was needed to improve the sensitivity and the limits of quantification of the MF-qPCR method to be similar to those of conventional qPCR. The MF-qPCR method was successfully demonstrated on untreated municipal wastewater, treated municipal wastewater, and drinking water samples. The treated municipal wastewater samples had higher concentrations of all genes compared to those in the drinking water samples. Similarly, the untreated municipal wastewater samples had higher concentrations for all but one of the targeted genes compared to those in the treated municipal wastewater samples. The MF-qPCR method established in this study provides highly accurate quantitative information about numerous ARGs and other genes from environmental samples.
中文翻译：
同时分析数十种抗生素抗性和重金属抗性基因的微流定量聚合酶链反应方法
这项研究开发，优化和证明了一种微流定量聚合酶链反应（MF-qPCR）方法，用于同时定量检测39个抗生素抗性基因（ARG），5个重金属抗性基因，3个编码三种不同整合素整合酶的基因以及16S rRNA基因（用于衡量细菌总生物量）。由于MF-qPCR的模板体积比常规qPCR小得多（几纳升），因此需要一个预扩增步骤来提高灵敏度，并且MF-qPCR方法的定量限与常规qPCR相似定量PCR。MF-qPCR方法已在未经处理的市政废水，经过处理的市政废水和饮用水样品中成功得到证明。与饮用水样品相比，处理过的市政废水样品中所有基因的浓度更高。类似地，与处理过的市政废水样品相比，未经处理的市政废水样品中除一种目标基因外的所有目标基因的浓度都较高。这项研究中建立的MF-qPCR方法可提供有关环境样品中众多ARG和其他基因的高精度定量信息。
更新日期：2017-12-21
中文翻译：
同时分析数十种抗生素抗性和重金属抗性基因的微流定量聚合酶链反应方法
这项研究开发，优化和证明了一种微流定量聚合酶链反应（MF-qPCR）方法，用于同时定量检测39个抗生素抗性基因（ARG），5个重金属抗性基因，3个编码三种不同整合素整合酶的基因以及16S rRNA基因（用于衡量细菌总生物量）。由于MF-qPCR的模板体积比常规qPCR小得多（几纳升），因此需要一个预扩增步骤来提高灵敏度，并且MF-qPCR方法的定量限与常规qPCR相似定量PCR。MF-qPCR方法已在未经处理的市政废水，经过处理的市政废水和饮用水样品中成功得到证明。与饮用水样品相比，处理过的市政废水样品中所有基因的浓度更高。类似地，与处理过的市政废水样品相比，未经处理的市政废水样品中除一种目标基因外的所有目标基因的浓度都较高。这项研究中建立的MF-qPCR方法可提供有关环境样品中众多ARG和其他基因的高精度定量信息。
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