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Nucleic Acid Quantitation with Log-Linear Response Hybridization Probe Sets.
ACS Sensors ( IF 8.2 ) Pub Date : 2020-06-01 , DOI: 10.1021/acssensors.0c00052 Lucia R Wu 1 , John Z Fang 1 , Dmitriy Khodakov 1 , David Yu Zhang 1, 2
ACS Sensors ( IF 8.2 ) Pub Date : 2020-06-01 , DOI: 10.1021/acssensors.0c00052 Lucia R Wu 1 , John Z Fang 1 , Dmitriy Khodakov 1 , David Yu Zhang 1, 2
Affiliation
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Concentrations of different nucleic acid species in biological samples span many orders of magnitude. A real-time polymerase chain reaction maps the concentration of a target nucleic acid sequence log–linearly into cycle threshold to enable quantitation with a wide dynamic range but suffers from enzymatic biases. Here, we present a general design for constructing hybridization probe sets with highly log–linear response curves to enable accurate enzyme-free quantitation across large ranges (more than 6 logs) of target DNA concentrations. The sensitivity of each component probe is accurately adjusted via formulation stoichiometry to reduce the standard error of target quantitation down to 7%. As a proof of concept, we show multiplexed quantitation of three microRNA species in total RNA of the human brain and liver.
中文翻译：
使用对数线性响应杂交探针组进行核酸定量。
生物样品中不同核酸种类的浓度跨越多个数量级。实时聚合酶链反应可将目标核酸序列的浓度线性对数映射到循环阈值，以实现宽动态范围的定量分析，但存在酶促偏差。在这里，我们提出了构建具有高对数线性响应曲线的杂交探针组的通用设计，以实现在目标DNA浓度的大范围（超过6个对数）范围内的准确无酶定量。可通过配方化学计量法精确调整每个组分探针的灵敏度，以将目标定量的标准误差降低至7％。作为概念的证明，我们显示了人脑和肝脏总RNA中三种microRNA的多重定量。
更新日期：2020-06-26
中文翻译：
使用对数线性响应杂交探针组进行核酸定量。
生物样品中不同核酸种类的浓度跨越多个数量级。实时聚合酶链反应可将目标核酸序列的浓度线性对数映射到循环阈值，以实现宽动态范围的定量分析，但存在酶促偏差。在这里，我们提出了构建具有高对数线性响应曲线的杂交探针组的通用设计，以实现在目标DNA浓度的大范围（超过6个对数）范围内的准确无酶定量。可通过配方化学计量法精确调整每个组分探针的灵敏度，以将目标定量的标准误差降低至7％。作为概念的证明，我们显示了人脑和肝脏总RNA中三种microRNA的多重定量。
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