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Related Concept Videos

PCR01:32

PCR

Overview
RACE - Rapid Amplification of cDNA Ends02:35

RACE - Rapid Amplification of cDNA Ends

Rapid Amplification of cDNA Ends, or RACE, is one of the most effective methods to obtain a full-length cDNA from an mRNA sequence between a known internal region to the unknown sequence at the 5’ or 3’ end. The unknown region is cloned in the cDNA by a gene-specific primer that binds the known end, and a hybrid primer that attaches a predefined anchor sequence to the unknown end of the cDNA. The sequence in between is amplified by PCR with an anchor primer and a gene-specific primer.
Since the...
PCR - Polymerase Chain Reaction01:32

PCR - Polymerase Chain Reaction

Overview
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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Updated: May 8, 2026

Polymerase Chain Reaction: Basic Protocol Plus Troubleshooting and Optimization Strategies
09:00

Polymerase Chain Reaction: Basic Protocol Plus Troubleshooting and Optimization Strategies

Published on: May 22, 2012

Explanatory chapter: PCR primer design.

Rubén Álvarez-Fernández1

  • 1Department of Plant Sciences, University of Cambridge, Cambridge, United Kingdom.

Methods in Enzymology
|September 10, 2013
PubMed
Summary
This summary is machine-generated.

This guide explains polymerase chain reaction (PCR) primer design, covering general guidelines and specific applications. It also recommends useful software tools for optimizing primer selection.

Keywords:
Homologous recombinationMultiplex PCRPCR Primer designPrimer lengthReal-time PCRReverse-transcription PCR (RT-PCR)Target cloning

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Area of Science:

  • Molecular Biology
  • Genetics

Background:

  • Polymerase Chain Reaction (PCR) is a fundamental technique in molecular biology.
  • Effective PCR relies heavily on well-designed primers for successful amplification.
  • Primer design is a critical step that influences PCR specificity and efficiency.

Purpose of the Study:

  • To provide a comprehensive guide on designing primers for Polymerase Chain Reaction (PCR).
  • To outline general principles and specific strategies for primer design.
  • To recommend software tools that aid in the primer design process.

Main Methods:

  • Discussion of general primer design guidelines.
  • Exploration of primer design considerations for various PCR applications.
  • Compilation of a list of recommended primer design software.

Main Results:

  • Established best practices for designing effective PCR primers.
  • Highlighted application-specific primer design considerations.
  • Provided a resource of relevant software tools for researchers.

Conclusions:

  • Proper primer design is essential for successful and reproducible PCR experiments.
  • Utilizing recommended guidelines and software can significantly improve primer selection.
  • This chapter serves as a valuable resource for researchers performing PCR.