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用于环境风险评估的商业实时PCR阵列的验证:在研究小家鼠肝脏中p,p´-滴滴伊毒性中的应用

Validation of commercial real-time PCR-arrays for environmental risk assessment: Application to the study of p,p´-DDE toxicity in Mus spretus mice liver.

作者信息

Morales-Prieto Noelia, Pueyo Carmen, Abril Nieves

机构信息

Department of Biochemistry and Molecular Biology, Building Severo Ochoa, 2nd Floor, Agrifood Campus of International Excellence (CeiA3-UCO), Campus de Rabanales, University of Cordoba, Spain.

出版信息

Environ Pollut. 2017 Nov;230:178-188. doi: 10.1016/j.envpol.2017.06.031. Epub 2017 Jun 23.

DOI:10.1016/j.envpol.2017.06.031
PMID:28651089
Abstract

Data on gene transcription profiles provide a comprehensive assessment of the toxic and defensive mechanisms that are triggered by pollutants. PCR-arrays have emerged as a reliable tool for analyzing the expression of a panel of relevant, pathway- or disease-focused genes under uniform cycling conditions. By using SYBR Green-optimized primer assays, it is possible to simultaneously amplify a sample with high specificity and amplification efficiencies. However, commercial PCR-arrays target a limited group of organisms, excluding most of those with environmental relevance, as is the case with Mus spretus mice. Our previous works with M. spretus showed a high sequence similarity between M. spretus and the model organism M. musculus allowing the use of commercial platforms with M. spretus. This work demonstrates the successful application of a commercial PCR-array designed for the model organism M. musculus to assess the biological effects caused by the organochlorine pesticide p,p´-DDE in a focused panel of stress-related genes in M. spretus mice. This cost-effective, easy-to-use platform detected quantitative gene profiling differences between M. spretus hepatic RNA samples and generated data highly concordant with those obtained by absolute qRT-PCR, the most sensitive method to quantify transcripts. This platform is also suitable for use in field studies with free-living M. spretus mice for routine environmental risk assessment. Our results provide a broad impression of the biological consequences of p,p´-DDE on the hepatic health of mice.

摘要

基因转录谱数据提供了对污染物引发的毒性和防御机制的全面评估。PCR阵列已成为一种可靠的工具,用于在统一循环条件下分析一组相关的、以途径或疾病为重点的基因的表达。通过使用SYBR Green优化引物检测,可以以高特异性和扩增效率同时扩增样品。然而,商业PCR阵列针对的生物种类有限,排除了大多数与环境相关的生物,小家鼠就是这样的例子。我们之前对小家鼠的研究表明,小家鼠与模式生物小鼠之间具有高度的序列相似性,这使得可以将商业平台用于小家鼠。这项工作展示了一种为模式生物小鼠设计的商业PCR阵列成功应用于评估有机氯农药p,p´-滴滴伊对小家鼠一组应激相关基因所造成的生物学效应。这个经济高效、易于使用的平台检测到了小家鼠肝脏RNA样本之间的定量基因谱差异,并生成了与通过绝对定量逆转录PCR(最灵敏的转录本定量方法)获得的数据高度一致的数据。该平台也适用于对自由生活的小家鼠进行野外研究,以进行常规环境风险评估。我们的结果广泛展示了p,p´-滴滴伊对小鼠肝脏健康的生物学影响。

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