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PCR

PCR

Real Time RT-PCR

Real Time RT-PCR

Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...

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Related Experiment Video

Updated: Mar 11, 2026

Single Cell Multiplex Reverse Transcription Polymerase Chain Reaction After Patch-clamp

Single Cell Multiplex Reverse Transcription Polymerase Chain Reaction After Patch-clamp

Published on: June 20, 2018

Multi-template polymerase chain reaction.

Elena Kalle¹, Mikael Kubista², Christopher Rensing³

¹Department of Forest Mycology and Plant Pathology, Swedish University of Agricultural Sciences, Allmas alle 5, 75007 Uppsala, Sweden.

Biomolecular Detection and Quantification

|November 30, 2016

Summary

This summary is machine-generated.

Rigorous validation is essential for Polymerase Chain Reaction (PCR) applications, especially with multi-template samples. Addressing artifacts and inhibitors ensures accurate results in complex biological analyses.

Keywords:

CDCE, constant denaturing capillary electrophoresis Chimera DGGE, denaturing gradient gel electrophoresis DHPLC, denaturing high-performance liquid chromatography HPLC, high-performance liquid chromatography Multi-template PCR PAAG, polyacrylamide gel SSCA, single strand conformation analysis T-RFLP, terminal restriction fragment length polymorphism TGGE, temperature gradient gel electrophoresis

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Last Updated: Mar 11, 2026

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Multiplex Detection of Bacteria in Complex Clinical and Environmental Samples using Oligonucleotide-coupled Fluorescent Microspheres

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Published on: October 23, 2011

Area of Science:

Molecular Biology
Genetics
Biotechnology

Background:

Polymerase Chain Reaction (PCR) is a vital tool in biological research.
Inconsistent standards in PCR applications lead to inaccurate and unreliable results.
Multi-template samples present unique challenges, including artifacts and biases.

Approach:

Emphasizes the critical need for rigorous method validation before adopting new PCR applications.
Highlights the complexities of analyzing multi-template samples due to varying template concentrations and homologous sequences.
Discusses the formation of chimeras and heteroduplexes in multi-template PCR.
Addresses issues of differential amplification efficiencies and competition for reaction components.
Explains how inhibitors can exacerbate existing problems and affect different templates unevenly.

Key Points:

Multi-template samples are prone to artifacts like chimeras and heteroduplexes.
Template competition and varying amplification efficiencies distort original ratios.
Inhibitors introduce further complexity, with potentially differential effects on templates.
Lack of standardized methods for monitoring inhibition in multi-template PCR is a significant gap.

Conclusions:

Rigorous validation is paramount for reliable PCR results, particularly with complex samples.
Standardized approaches are needed to monitor inhibitory effects in multi-template PCR.
Ensuring compatibility between samples is crucial for accurate downstream analysis.