Reverse transcriptases Reversible reactions

The amplification of cDNA by combining a reverse transcriptase (RT) reaction with PCR is known as reverse transcription polymerase chain reaction (RT-PCR). A first-strand of cDNA is generated from RNA with RT in the presence of dNTP, Mg + by the use of either gene-specific RT primers or random hexamer primer at 50°C for 1 h. The reaction mixture is heated (e.g. 70°C for 15 min) and cooled. The resulting gene-specific RT reaction mixture is column purified and subjected to PCR amplification of cDNA. 

In the realm of natural product synthesis, Kepler and Rehder utilized the K-R reaction to synthesize ( )-calanolide A (56), a potent non-nucleosidal human irmnunodeficiency virus (HIV-1) specific reverse transcriptase inhibitor. Propiophenone 57 was allowed to react with acetic anhydride in the presence of sodium acetate to afford benzopyranone 58 in 56% yield subsequent deacetylation of 58 gave 59. Flavone 59 was then transformed to ( ) calanolide A (56) over several steps. 

This acronym stands for Reverse Transcriptase Polymerase Chain Reaction, a method used to first copy a strand of RNA into cDNA, then amplify it through standard PCR methods. 

RT-PCR reverse transcriptase polymerase chain reaction RTV ritonavir... 

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. 

This step removes any residual proteins. It is important to avoid contamination of phenol in the extracted phase because this will affect the reverse transcriptase reaction. It is also important to avoid the interphase, which is enriched with DNA. 

Abrahamsen HN, Steiniche T, Nexo E, et al. Towards quantitative mRNA analysis in paraffin-embedded tissues using real-time reverse transcriptase-polymerase chain reaction. A methodological study on lymph nodes from melanoma patients. J. Mol. Diagn. 2003 5 34-41. 

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. 

Mangham DC, Williams A, McMullan DJ, et al. Ewing s sarcoma of bone the detection of specific transcripts in a large, consecutive series of formalin-fixed, decalcified, paraffin-embedded tissue samples using the reverse transcriptase -polymerase chain reaction. Histopathology 2006 48 363-376. 

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