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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...

PCR

PCR

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qPCRTag Analysis - A High Throughput, Real Time PCR Assay for Sc2.0 Genotyping

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Introduction to kinetic (real-time) PCR.

¹Applied Science, Roche Diagnostics, Mt Wellington, Auckland, New Zealand.

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

|March 3, 2007

Summary

Real-time PCR, also known as kinetic PCR, offers speed and reduced contamination risk, making it a superior alternative to conventional PCR. This technique is rapidly evolving with new applications emerging constantly.

Area of Science:

Molecular Biology
Biotechnology

Background:

Kinetic PCR (real-time PCR) has undergone rapid development since the early 1990s.
It is increasingly replacing conventional PCR due to advantages like speed, reduced hands-on time, and closed-tube format minimizing contamination.

Purpose of the Study:

To cover the fundamental principles of kinetic PCR.
To discuss critical pre-amplification parameters for successful real-time PCR assays.

Main Methods:

Explanation of the basis of kinetic PCR.
Discussion of nucleic acid extraction protocols.
Overview of complementary DNA synthesis for gene expression studies.

Main Results:

Real-time PCR offers significant improvements over conventional PCR.

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Careful consideration of pre-amplification steps is crucial for assay performance.

Conclusions:

Kinetic PCR is a rapidly advancing technique with broad applicability.
Optimizing nucleic acid extraction and cDNA synthesis is essential for gene expression analysis using real-time PCR.