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Related Concept Videos

Spontaneous and Induced Mutations01:30

Spontaneous and Induced Mutations

Spontaneous mutations arise infrequently during DNA replication due to errors in the process. A key factor behind these errors is tautomeric shifts in nitrogenous bases, where bases transition from keto to enol forms or amino to imino forms. This shift can alter base-pairing rules, leading to mutations. Additionally, reactive oxygen species (ROS) arising from aerobic metabolism can damage DNA, resulting in depurination (loss of a purine base) or depyrimidination (loss of a pyrimidine base).
Mismatch Repair01:36

Mismatch Repair

Overview
Mismatch Repair01:20

Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the new strand and replacing the erroneous bases with correct nucleotides. Mismatch repair is coordinated by many proteins in both prokaryotes and eukaryotes.
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
In vitro Mutagenesis01:16

In vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.
Proofreading01:31

Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore,  it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Proofreading01:43

Proofreading

Synthesis of new DNA molecules starts when DNA polymerase links nucleotides together in a sequence that is complementary to the template DNA strand. DNA polymerase has a higher affinity for the correct base to ensure fidelity in DNA replication. The DNA polymerase furthermore proofreads during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.Errors during Replication Are Corrected by the DNA Polymerase EnzymeGenomic DNA is synthesized in...

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Related Experiment Video

Updated: Jun 10, 2026

Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast
07:18

Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast

Published on: May 15, 2018

Mutagen: a random mutagenesis method providing a complementary diversity generated by human error-prone DNA

Philippe Mondon1, David Grand, Nathalie Souyris

  • 1Antibody Engineering and Molecular Evolution Department, MilleGen SA, Labège, France. philippe.mondon@millegen.com

Methods in Molecular Biology (Clifton, N.J.)
|August 3, 2010
PubMed
Summary

This study introduces MutaGen, a novel random mutagenesis technique using error-prone human DNA polymerases. MutaGen efficiently generates diverse gene and protein variant libraries for directed evolution without needing prior structural data.

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Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
09:01

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli

Published on: March 16, 2011

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Last Updated: Jun 10, 2026

Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast
07:18

Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast

Published on: May 15, 2018

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
09:01

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli

Published on: March 16, 2011

Area of Science:

  • Molecular Biology
  • Biotechnology
  • Protein Engineering

Background:

  • Random mutagenesis is crucial for generating protein variant libraries for directed evolution.
  • Existing methods may require specific structural or mechanistic information.
  • Discovering beneficial mutations often relies on unbiased exploration.

Purpose of the Study:

  • To introduce a novel random mutagenesis method, MutaGen, utilizing human error-prone DNA polymerases.
  • To enable efficient gene and protein randomization through a simplified process.
  • To generate diverse variant libraries with complementary mutational profiles.

Main Methods:

  • Employed human DNA polymerases (pol beta, pol eta, pol iota) for random mutagenesis.
  • Utilized a single replication step followed by selective PCR amplification.
  • Characterized mutation rates and mutational spectra of generated libraries.

Main Results:

  • MutaGen successfully generated variant libraries with varying mutation rates.
  • The method produced complementary mutational spectra, enhancing library diversity.
  • Leveraged the inherent mutation bias of error-prone polymerases effectively.

Conclusions:

  • MutaGen offers a powerful and versatile tool for gene and protein randomization.
  • The method simplifies library generation for directed protein evolution.
  • It provides an alternative approach that does not require prior structural or mechanistic insights.