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 ...

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...

The development of microfluidic tools for high-throughput nucleic acid analysis has become a burgeoning area of research in the post-genome era. Here, we have developed a microfluidic chip to perform 72 parallel 450-pL RT-PCRs. We took advantage of Taqman hydrolysis probe chemistry to detect RNA templates as low as 34 copies. The device and method presented here may enable highly parallel singl...

Reverse transcriptase polymerase chain reaction (RT-PCR) has become a well-established and powerful molecular technique for studying ribonucleic acids. It is used in medical diagnostics for detecting viral RNA, in hematology and oncology for detecting chimeric transcripts of rearranged genes (1), and in the broad area of research applications in gene expression studies. Since its introduction (...