Mitchell Kendall, Iadarola Michael J
Neurobiology and Pain Therapeutics Section, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA.
Methods Mol Biol. 2010;617:279-95. doi: 10.1007/978-1-60327-323-7_21.
Frequently, it is important to ascertain whether a molecule that is involved in one model of pain is also involved in other models of pain. Similarly, it may be important to determine whether a molecule involved in nociception in one tissue is also expressed in other tissues and to ascertain the degree of enrichment. Additionally, before initiating a complex set of experiments or purchasing an expensive immunoassay kit, it may be useful to obtain initial supporting evidence to justify the time and money. Is the transcript for the target receptor, protein, or peptide precursor present in, for example, the dorsal root ganglion? And, if present, how abundant is it? Here is where the power of PCR can be applied to obtain a quick but informative answer. In this chapter, we mainly detail the use of gel-based RT-PCR and also provide suggestions on tissue dissection and interpretation of results. The use of gel-based RT-PCR can address many of the questions of abundance or tissue specificity with a minimum of expense and time.
通常，确定参与一种疼痛模型的分子是否也参与其他疼痛模型很重要。同样，确定一种在一个组织中参与伤害感受的分子是否也在其他组织中表达并确定其富集程度可能也很重要。此外，在启动一系列复杂的实验或购买昂贵的免疫分析试剂盒之前，获得初步的支持证据以证明时间和金钱的合理性可能会很有用。例如，背根神经节中是否存在目标受体、蛋白质或肽前体的转录本？如果存在，其丰度如何？这就是PCR的强大功能可以用来快速获得有用答案的地方。在本章中，我们主要详细介绍基于凝胶的RT-PCR的使用，并提供关于组织解剖和结果解释的建议。基于凝胶的RT-PCR的使用可以用最少的费用和时间解决许多关于丰度或组织特异性的问题。
