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首页> 外文期刊>Brain research. Brain research protocols >Novel quantitative reverse-transcribed polymerase chain reaction of mu opioid receptor mRNA level.
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Novel quantitative reverse-transcribed polymerase chain reaction of mu opioid receptor mRNA level.

机译：新型定量逆转录聚合酶链反应μ阿片受体mRNA水平。

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摘要

Reverse-transcribed polymerase chain reaction (RT-PCR) can quantify gene transcripts at low levels and in small samples. Semi-quantitative and quantitative RT-PCR has significant advantages over traditional RNA assays, such as Northern blotting and ribonuclease protection assay. However, owing to the exponential nature of PCR, considerable effort is required to verify linearity of the reaction. Thus, care must be taken to detect small but physiologically relevant changes in gene expression. Using a rapid and highly sensitive RT-PCR method, TaqMan real-time RT-PCR, we determined agonist-induced changes in rat mu opioid receptor (MOR) mRNA levels in cultured cells and compared our results with those obtained by radiolabeled quantitative RT-PCR, which is also highly sensitive but much more time-consuming than TaqMan RT-PCR. Both methods showed up-regulation of agonist-induced MOR. TaqMan RT-PCR showed a similar sensitivity to radiolabeled quantitative RT-PCR and is suitable for the measurement of large numbers of samples. Moreover, no need for radiolabeled compounds is also an advantage of TaqMan PCR. This protocol will probably be useful for quantifying MOR in animal and human tissues.
机译：逆转录聚合酶链反应（RT-PCR）可以定量检测低含量和少量样品中的基因转录物。半定量和定量RT-PCR与传统的RNA分析（例如Northern印迹和核糖核酸酶保护分析）相比具有显着优势。然而，由于PCR的指数性质，需要大量的努力来验证反应的线性。因此，必须注意检测基因表达中微小但生理相关的变化。使用快速且高度灵敏的RT-PCR方法，TaqMan实时RT-PCR，我们确定了激动剂诱导的培养细胞中大鼠阿片类阿片受体（MOR）mRNA水平的变化，并将我们的结果与通过放射性标记的定量RT-PCR获得的结果进行了比较PCR，它也是高度敏感的，但比TaqMan RT-PCR耗时得多。两种方法均显示出激动剂诱导的MOR上调。 TaqMan RT-PCR显示出与放射性标记的定量RT-PCR相似的灵敏度，适用于测量大量样品。而且，不需要放射性标记的化合物也是TaqMan PCR的优势。该协议可能对定量动物和人类组织中的MOR有帮助。

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