Quantitative Real-Time PCR Analysis of Degradome Gene Expression
Dissection of the contribution of proteases and inhibitors in the complex molecular events involved in cancer initiation, growth, and spread requires as a starting point detailed knowledge of the degradome genes that are expressed and dysregulated in cancer. This information identifies candidate genes for functional investigations and also reveals potential markers of disease progression and severity. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis provides optimal sensitivity and specificity for analysis of RNA from human tumors and nonneoplastic tissues. In this chapter, we outline basic qRT-PCR technologies, and approaches for normalization and analysis of expression data. Degradation of RNA is a major problem for microarray analyses, but we demonstrate that TaqMan® qRT-PCR is a remarkably robust technique that can provide reliable information on archival specimens that would not be appropriate for other transcriptomic analyses. We also highlight the utility of low-density TaqMan arrays for degradome expression analysis.
KeywordsUrothelial Carcinoma Taqman Probe Reverse Transcription Reaction Molecular Beacon Standard Curve Method
Unable to display preview. Download preview PDF.