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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.
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.
Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
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).

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Related Experiment Video

Updated: Jun 5, 2026

Site-Directed Mutagenesis for In Vitro and In Vivo Experiments Exemplified with RNA Interactions in Escherichia Coli
07:04

Site-Directed Mutagenesis for In Vitro and In Vivo Experiments Exemplified with RNA Interactions in Escherichia Coli

Published on: February 5, 2019

Site-directed mutagenesis.

Patricia E Carrigan1, Petek Ballar, Sukru Tuzmen

  • 1Biodesign Institute, Arizona State University, Phoenix, AZ, USA.

Methods in Molecular Biology (Clifton, N.J.)
|January 5, 2011
PubMed
Summary
This summary is machine-generated.

Site-directed mutagenesis enables precise gene and protein modification for studying structure-function relationships. This versatile technique, utilizing synthetic oligonucleotides, is essential for disease gene characterization.

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Last Updated: Jun 5, 2026

Site-Directed Mutagenesis for In Vitro and In Vivo Experiments Exemplified with RNA Interactions in Escherichia Coli
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Published on: February 5, 2019

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Site-directed mutagenesis is a cornerstone technique in molecular biology.
  • It has been instrumental in characterizing gene and protein structure-function relationships for over 30 years.
  • Understanding protein interactions, binding domains, and enzyme active sites relies on this method.

Purpose of the Study:

  • To provide a detailed overview of site-directed mutagenesis.
  • To highlight the essential steps and considerations for successful implementation.
  • To underscore the technique's versatility and applicability in research.

Main Methods:

  • Experimental alteration of nucleotide sequences using synthetic oligonucleotides.
  • Employing complementary oligonucleotides with internal mismatches to direct mutations.
  • Application for single point mutations, multiple mutations, insertions, and deletions.

Main Results:

  • Site-directed mutagenesis allows for targeted modifications of DNA sequences.
  • The technique is adaptable for various genetic alterations, including point mutations, insertions, and deletions.
  • Commercial kits have streamlined the process, making it quick, reliable, and accessible.

Conclusions:

  • Site-directed mutagenesis is a powerful and versatile tool for investigating gene and protein functions.
  • Its broad applicability, particularly in disease gene characterization, makes it indispensable.
  • The technique's reliability and commercial availability facilitate its widespread use in research settings.