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Related Concept Videos

Real Time RT-PCR02:57

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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Quantitative real-time PCR-based analysis of gene expression.

J Jozefczuk1, James Adjaye

  • 1Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics (Molecular Embryology and Aging group), Ihnestrasse 63-73, Berlin, Germany.

Methods in Enzymology
|September 28, 2011
PubMed
Summary

Quantitative real-time polymerase chain reaction (QRT-PCR) is a versatile gene expression analysis tool. This guide details QRT-PCR principles for accurate mRNA quantification in various biological samples.

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • Quantitative real-time polymerase chain reaction (QRT-PCR) is a widely adopted technique.
  • It is crucial for quantitative gene expression analysis in research, medicine, and diagnostics.
  • QRT-PCR is used to analyze messenger RNA (mRNA) expression patterns and compare mRNA levels across samples.

Purpose of the Study:

  • To provide a comprehensive step-by-step guide to QRT-PCR principles.
  • To elucidate the foundational concepts for successful QRT-PCR experiments.
  • To highlight the utility of QRT-PCR in various scientific domains.

Main Methods:

  • Detailed explanation of QRT-PCR methodology.
  • Focus on principles for accurate mRNA quantification.
  • Guidance on experimental design and execution.

Main Results:

  • QRT-PCR offers a large dynamic range, high sensitivity, and sequence-specificity.
  • It enables comparisons between normal and abnormal tissues.
  • It serves as independent validation for microarray and next-generation sequencing (NGS) results.

Conclusions:

  • QRT-PCR is an indispensable tool for precise gene expression analysis.
  • Understanding QRT-PCR principles is key to reliable experimental outcomes.
  • The technique supports diverse applications from basic research to clinical diagnostics.