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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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Updated: Mar 11, 2026

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

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Published on: June 20, 2018

Multiplex Analyses Using Real-Time Quantitative PCR.

Steve F C Hawkins¹, Paul C Guest²

¹Bioline Reagents Limited, Unit 16, The Edge Business Centre, Humber Road, London, NW2 6EW, UK. steve.hawkins@bioline.com.

Methods in Molecular Biology (Clifton, N.J.)

|November 30, 2016

Summary

This summary is machine-generated.

Multiplex quantitative polymerase chain reaction (qPCR) enables simultaneous gene expression analysis. This method efficiently quantifies up to five targets in real time using probe-based assays.

Keywords:

Amplicon Fluorescent dyes Multiplex analysis Quantitation Taq polymerase cDNA mRNA qPCR

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Area of Science:

Molecular Biology
Genetics
Biochemistry

Background:

Quantitative polymerase chain reaction (qPCR) is a standard technique for real-time gene expression detection and quantification.
Multiplex qPCR utilizes probe-based assays with unique fluorescent dyes for simultaneous detection of multiple targets.
This multiplexing capability allows for rapid measurement of several genes of interest within a single reaction.

Purpose of the Study:

To describe a method for simultaneous real-time quantitation of multiple amplicons using qPCR.
To demonstrate the utility of SensiFAST and SensiFAST One-Step probe kits for multiplex qPCR applications.

Main Methods:

Utilized probe-based assays with unique fluorescent dyes for multiplexing.
Employed SensiFAST and SensiFAST One-Step probe kits for the simultaneous quantitation of gene expression.
Performed real-time quantitative polymerase chain reaction to detect and quantify amplicons.

Main Results:

Successfully achieved simultaneous real-time quantitation of up to 5 amplicons in a single reaction.
Demonstrated the effectiveness of the described method for multiplex qPCR.
Validated the performance of SensiFAST and SensiFAST One-Step probe kits in multiplex assays.

Conclusions:

The described method enables efficient and rapid multiplex quantitative polymerase chain reaction.
SensiFAST and SensiFAST One-Step probe kits are suitable for simultaneous real-time quantitation of multiple targets.
This approach facilitates high-throughput gene expression analysis.