Reverse transcriptase polymerase chain reaction and

The methods used for the evaluation of regulation of gene expression are too numerous to be described in detail here. They include Northern analysis to determine levels of a particular mRNA, nuclear run on to determine whether an increase in mRNA is due to an increase in the rate of transcription, and promoter deletion analysis to identify specific elements in the promoter region responsible for the control of expression. Of much current interest is the use of microarrays that permit the study of the expression of hundreds to thousands of genes at the same time. Reverse transcriptase-polymerase chain reaction and RNase protection assay techniques are used to amplify and quantitate mRNAs, while the electrophoretic mobility shift assay is used to measure binding of a transcription factor to its specific DNA consensus sequence. 

Kohler, R., Degenhardt, C., Kuhn, M., Runkel, N., Paul, M., and Hoyer, J. 2000. Expression and function of endothelial Ca2+-activated K+ channels in human mesenteric artery A single-cell reverse transcriptase-polymerase chain reaction and electrophysiological study in situ. Circ. Res. 87 496-503. 

Kodera, Y., Nakanishi, H., Yamamura, Y., Shimizu, Y., Torii, A., Hirai, T., et al.. Prognostic value and clinical implications of disseminated cancer cells in the peritoneal cavity detected by reverse transcriptase-polymerase chain reaction and cytology. Int. J. Cancer 79, 429-433 (1998). 

O., et al. Comparative detection of lymph node micrometastases of stage II colorectal cancer by reverse transcriptase polymerase chain reaction and immunohistochemistry. 

Inducible nitric oxide synthase (iNOS EC 1.14.13.39) mRNA in bronchoalvelar lavage cells (using reverse transcriptase-polymerase chain reaction) and the magnitude of NO-dependent chemiluminescence from alveolar macrophages correlated with the pathology of silica-exposed coal miners chest radiographs (Castranova et al. 1998). 

Dunne, A. L., and Crowe, S. M. (1997). Comparison of branched DNA and reverse transcriptase polymerase chain reaction for quantifying six different HTV-1 subtypes in plasma. AIDS 11,2-3. 

Cronin M, Pho M, Dutta D, et al. Measurement of gene expression in archival paraffin-embedded tissues development and performance of a 92-gene reverse transcriptase-polymerase chain reaction assay. Am. J. Pathol. 2004 164 35—42. 

The RNA molecules, ribosomal RNA (rRNA) and messenger RNA (mRNA) play key roles in the protein synthesis. The amount of RNA in individual cells or in a community may, therefore, be taken as an indicator of protein synthesis and, thus, microbial activity. The number of active cells can be detected by fluorescent in situ hybridization (FISH) (Amann et al. 1995). By this method, individual cells carrying high concentrations of rRNA, situated on ribosomes, are quantified by fluorescence microscopy. The amount of rRNA in a community can also be detected by Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), where rRNA extracted from soil is detected by creating a DNA copy and separating by gel electrophoresis (Duineveld et al. 2001). 

Regarding reverse transcriptase polymerase chain reaction (RT-PCR) analysis to assess a splice site variant, as seen in Fig. 12.6, if one designs forward (F-) and reverse (R-) RT-PCR primers to span the SNP (which in turn creates or abolishes a splice site), one wiU have different size PCR products (labeled a, b, and c in the figure) that can easily be resolved on a gel. 

Chu, K. H., Wong, S. H., and Leung, P. S. C. (2000). Tropomyosin is the major mollusk allergen Reverse transcriptase polymerase chain reaction, expression and IgE reactivity. Mar. Biotechnol. 2,499-509. 

E. Therapeutic response Efficacy of Infergen therapy was determined by measurement of serum alanine aminotransferase (ALT) concentrations at the end of therapy (24 weeks) and following 24 weeks of observation after the end of treatment of adults with chronic HCV infection. Serum HCV RNA was also assessed using a quantitative reverse transcriptase polymerase chain reaction (RT-PCR). At the end of 24 weeks of treatment, ALT normalization was observed in 39% of patients on Infergen and in 35% of patients on interferon alfa-2b Intron A). Only 17% of patients in each group... 

Heniford, B W., Shum-Siu, A, Leonberger, M., and Hendler, F J (1993) Variation m cellular EGF receptor mRNA expression demonstrated by in situ reverse transcriptase polymerase chain reaction. Nucleic Acids Res 21,3159-3166... 

Raman spectroscopy can offer a number of advantages over traditional cell or tissue analysis techniques used in the field of TE (Table 18.1). Commonly used analytical techniques in TE include the determination of a specific enzyme activity (e.g. lactate dehydrogenase, alkaline phosphatase), the expression of genes (e.g. real-time reverse transcriptase polymerase chain reaction) or proteins (e.g. immunohistochemistry, immunocytochemistry, flow cytometry) relevant to cell behaviour and tissue formation. These techniques require invasive processing steps (enzyme treatment, chemical fixation and/or the use of colorimetric or fluorescent labels) which consequently render these techniques unsuitable for studying live cell culture systems in vitro. Raman spectroscopy can, however, be performed directly on cells/tissue constructs without labels, contrast agents or other sample preparation techniques. 

RNA Extraction and Reverse Transcriptase Polymerase Chain Reaction... 

Endrizzi K, Fischer J, Klein K, Schwab M, Nussler A, Neuhaus P, Eichelbaum M, Zanger UM. Discriminative quantification of cytochrom P4502D6 and 2D7/8 psuedogene expression by TaqMan real-time reverse transcriptase polymerase chain reaction. Anal Biochem 2002 15 121-131. 

Wesselingh, S. L., Takahashi, K., Glass,J. D., McArthur,J. C., Griffin,J. W., and Griffin, D. E. (1997). Cellular localization of tumor necrosis factor mRNA in neurological tissue from HIV-infected patients by combined reverse transcriptase/polymerase chain reaction in situ hybridization and immunohistochemistry. J. Neuroimmunol. 74, 1—8. 

---