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首页> 外文期刊>Journal of Cellular Physiology >Evaluation and identification of reliable reference genes for pharmacogenomics, toxicogenomics, and small RNA expression analysis.
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Evaluation and identification of reliable reference genes for pharmacogenomics, toxicogenomics, and small RNA expression analysis.

机译：评估和可靠的识别参考基因药物基因组学,toxicogenomics和小RNA表达分析。

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Pharmacogenomics, toxicogenomics, and small RNA expression analysis are three of the most active research topics in the biological, biomedical, pharmaceutical, and toxicological fields. All of these studies are based on gene expression analysis, which requires reference genes to reduce the variations derived from different amounts of starting materials and different efficiencies of RNA extraction and cDNA synthesis. Thus, accurate normalization to one or several constitutively expressed reference genes is a prerequisite to valid gene expression studies. Although selection of reliable reference genes has been conducted in previous studies in several animals and plants, no research has been focused on pharmacological targets, and very few studies have had a toxicological context. More interestingly, no studies have been performed to identify reference genes for small RNA analysis although small RNA, particularly microRNA (miRNA)-related research is currently one of the fastest-moving topics. In this study, using MCF-7 breast cancer cells as a model, we employed quantitative real-time PCR (qRT-PCR), one of the most reliable methods for gene expression analysis in many research fields, to evaluate and to determine the most reliable reference genes for pharmacogenomics and toxicogenomics studies as well as for small RNA expression analysis. We tested the transcriptional expression of five protein-coding genes as well as five non-coding genes in MCF-7 cells treated with five different pharmaceuticals or toxicants [paclitaxel (PTX), gossypol (GOS), methyl jasmonate (JAS), L-nicotine (NIC), and melamine (mela)] and analyzed the stability of the selected reference genes by four different methods: geNorm, NormFinder, BestKeeper, and the comparative DeltaCt method. According to our analysis, a protein-coding gene, hTBCA and four non-coding genes, hRNU44, hRNU48, hU6, and hRNU47, appear to be the most reliable reference genes for the five chemical treatments. Similar results were also obtained in dose-response and time-course assays with gossypol (GOS) treatment. Our results demonstrated that traditionally used reference genes, such as 18s RNA, beta-actin, and GAPDH, are not reliable reference genes for pharmacogenomics and toxicogenomics studies. In contrast, hTBCA and small RNAs are more stable during drug treatment, and they are better reference genes for pharmacogenomics and toxicogenomics studies. To widely use these genes as reference genes, these results should be corroborated by studies with other human cell lines and additional drugs classes and hormonal treatments.
机译：药物基因组学、toxicogenomics和小RNA三个最活跃的表达分析研究主题在生物、生物医学、制药、和毒理学领域。这些研究都是基于基因表达分析,这就需要参考基因减少来自不同的变化大量的原料和不同效率的RNA提取和互补合成。一些既定的参考基因表达是一个有效的基因表达的前提研究。在以往的研究的基因一些动物和植物,没有研究集中在药理指标,并很少有毒性的研究背景。有趣的是,没有进行研究确定参考小RNA基因分析虽然小RNA,特别是微(microrna) -相关研究是目前的一个当地的话题。乳腺癌细胞模型,我们使用实时定量PCR(存在),其中的一个基因表达的最可靠的方法在许多研究领域分析,评估和以确定最可靠的参考基因药物基因组学与toxicogenomics研究以及对小RNA表达分析。测试五的转录表达蛋白质编码基因以及五个非编码基因MCF-7细胞五个不同的对待药物或毒物(紫杉醇(PTX),棉子酚(GOS),甲基jasmonate(雅),L-nicotine (NIC),三聚氰胺(梅拉)]稳定性的分析选择的参考基因通过四种不同的方法:geNorm,NormFinder、BestKeeper和比较DeltaCt方法。蛋白质编码基因,hTBCA和四个非编码五个基因是最可靠的参考化学治疗。获得的剂量效应和时间进程化验棉子酚(GOS)治疗。证明了传统参考基因,如18 s RNA, beta-actin GAPDH,不可靠的参考基因药物基因组学和toxicogenomics研究。相反,hTBCA和小分子rna更稳定在药物治疗,他们是更好的参考药物基因组学和基因toxicogenomics研究。参考基因,这些结果与其他人类细胞通过这方面的研究行和额外的类和激素的药物治疗方法。

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