Abstract
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.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Yang HY, Mitchell K, Keller JM, Iadarola MJ (2007) Peripheral inflammation increases Scya2 expression in sensory ganglia and cytokine and endothelial related gene expression in inflamed tissue. J Neurochem 103:1628–1643
Coghill RC, McHaffie JG, Yen YF (2003) Neural correlates of interindividual differences in the subjective experience of pain. Proc Natl Acad Sci U S A 100:8538–8542
Caterina MJ, Schumacher MA, Tominaga M, Rosen TA, Levine JD, Julius D (1997) The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature 389:816–824
Olah Z, Szabo T, Karai L, Hough C, Fields RD, Caudle RM, Blumberg PM, Iadarola MJ (2001) Ligand-induced dynamic membrane changes and cell deletion conferred by vanilloid receptor 1. J Biol Chem 276:11021–11030
Karai L, Brown DC, Mannes AJ, Connelly ST, Brown J, Gandal M, Wellisch OM, Neubert JK, Olah Z, Iadarola MJ (2004) Deletion of vanilloid receptor 1-expressing primary afferent neurons for pain control. J Clin Invest 113:1344–1352
Brown DC, Iadarola MJ, Perkowski SZ, Erin H, Shofer F, Laszlo KJ, Olah Z, Mannes AJ (2005) Physiologic and antinociceptive effects of intrathecal resiniferatoxin in a canine bone cancer model. Anesthesiology 103:1052–1059
Gavva NR (2008) Body-temperature maintenance as the predominant function of the vanilloid receptor TRPV1. Trends Pharmacol Sci 29:550–557
Mitchell K, Yang HY, Berk JD, Tran JH, Iadarola MJ (2009) Monocyte chemoattractant protein-1 in the choroid plexus: a potential link between vascular pro-inflammatory mediators and the CNS during peripheral tissue inflammation. Neuroscience 158:885–895
Ali-Seyed M, Laycock N, Karanam S, Xiao W, Blair ET, Moreno CS (2006) Cross-platform expression profiling demonstrates that SV40 small tumor antigen activates Notch, Hedgehog, and Wnt signaling in human cells. BMC Cancer 6:54
Brody JR, Calhoun ES, Gallmeier E, Creavalle TD, Kern SE (2004) Ultra-fast high-resolution agarose electrophoresis of DNA and RNA using low-molarity conductive media. Biotechniques 37:598 600, 602
Yang HY, Wilkening S, Iadarola MJ (2001) Spinal cord genes enriched in rat dorsal horn and induced by noxious stimulation identified by subtraction cloning and differential hybridization. Neuroscience 103:493–502
Chakravarti B, Louie M, Ratanaprayul W, Raval A, Gallagher S, Chakravarti DN (2008) A highly uniform UV transillumination imaging system for quantitative analysis of nucleic acids and proteins. Proteomics 8:1789–1797
Mitchell K, Yang HY, Tessier PA, Muhly WT, Swaim WD, Szalayova I, Keller JM, Mezey E, Iadarola MJ (2008) Localization of S100A8 and S100A9 expressing neutrophils to spinal cord during peripheral tissue inflammation. Pain 134:216–231
Kim AY, Tang Z, Liu Q, Patel KN, Maag D, Geng Y, Dong X (2008) Pirt, a phosphoinositide-binding protein, functions as a regulatory subunit of TRPV1. Cell 133:475–485
Acknowledgments
This research was supported by the Intramural Research Program, NIDCR, NIH, DHHS. We thank Dr. H.-Y. T. Yang for helpful comments.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2010 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Mitchell, K., Iadarola, M.J. (2010). RT-PCR Analysis of Pain Genes: Use of Gel-Based RT-PCR for Studying Induced and Tissue-Enriched Gene Expression. In: Szallasi, A. (eds) Analgesia. Methods in Molecular Biology, vol 617. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60327-323-7_21
Download citation
DOI: https://doi.org/10.1007/978-1-60327-323-7_21
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-60327-322-0
Online ISBN: 978-1-60327-323-7
eBook Packages: Springer Protocols