INTRODUCTION The first step in competitive RT-PCR is the synthesis and purification of the synthetic competitor. This is an RNA molecule designed to be reverse-transcribed and PCR-amplified with the same efficiency as the endogenous transcript of interest. Once the competitor molecule has been prepared, as described in this protocol, competitive PCR can be carried out, as described in Comptetit...

Various approaches can now be taken for amplification of RNA transcripts using the polymerase chain reaction (PCR). Here, we compare three such methods: (i) uncoupled reverse transcription (RT)-PCR (using separate reactions for cDNA synthesis and PCR), (ii) continuous RT-PCR (in which RT and DNA amplification occur in an uninterrupted reaction) using either a single enzyme for both RT and DNA a...

Reverse transcription PCR (RT-PCR) represents a sensitive and powerful tool for analyzing RNA. While it has tremendous potential for quantitative applications, a comprehensive knowledge of its technical aspects is required. Successful quantitative RT-PCR involves correction for experimental variations in individual RT and PCR efficiencies. This review addresses the mathematics of RT-PCR, choice...

BACKGROUND Real-time polymerase chain reaction (PCR) utilizing the LightCycler and similar systems is an increasingly used technique for quantitative reverse transcription (RT)-PCR of mRNA levels from genes of immunologic interest. A commonly encountered limitation with these systems is that the fluorescence induced by SYBR Green (a fluorophore that binds double-stranded DNA) can result from pr...

Preoperative peripheral blood reverse transcription-PCR (RT-PCR) for prostate-specific antigen (PSA) [RT-PCR-PSA] is not associated with an increased risk of progression after radical prostatectomy. We tested the hypothesis that early postoperative peripheral blood RT-PCR-PSA would detect prostate cancer cells persisting in the circulation that would be associated with disease progression. The ...

Ga2O3/Ag/Ga2O3-laminated films with high electrical conductivity and ultraviolet (UV) transparency were achieved by radio frequency magnetron sputtering at room temperature (RT) on quartz glass. The influence of annealing ambient the structural, optical properties investigated in detail. As increases, bandgap Ga2O3-laminated widens. exhibited good photoelectric performance a figure-of-merit (FO...

PCR is widely used for the amplification of DNA and reverse-transcribed RNA. In recent years, real-time PCR has been introduced as a new technique for mRNA quantitation. However, because of the high cost of real-time PCR equipment and supplies, especially using custom-made probes, quantitative RT-PCR is a valuable alternative method. Several quantitative PCR protocols describe the use of a sing...