Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

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...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Life and Times in Engineering and Chemical Engineering.

Annual review of chemical and biomolecular engineering·2020
Same author

Genetic modification of mouse bone marrow by lentiviral vector-mediated delivery of hypoxanthine-Guanine phosphoribosyltransferase short hairpin RNA confers chemoprotection against 6-thioguanine cytotoxicity.

Transplantation proceedings·2013
Same author

Genotoxicity profiles of common alkyl halides and esters with alkylating activity.

Mutation research·2007
Same author

Time-of-flight variant to image mixing of granular media in a 3D fluidized bed.

Journal of magnetic resonance (San Diego, Calif. : 1997)·2007
Same author

Rapid two-dimensional imaging of bubbles and slugs in a three-dimensional, gas-solid, two-phase flow system using ultrafast magnetic resonance.

Physical review. E, Statistical, nonlinear, and soft matter physics·2007
Same author

Real-time measurement of bubbling phenomena in a three-dimensional gas-fluidized bed using ultrafast magnetic resonance imaging.

Physical review letters·2006

Related Experiment Video

Updated: Jul 24, 2026

Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
09:52

Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii

Published on: July 12, 2013

Mis-targeting of multiple gene disruption constructs containing hisG.

J F Davidson1, R H Schiestl

  • 1Department of Cancer Cell Biology, Harvard School of Public Health, Boston, MA 02115, USA.

Current Genetics
|December 29, 2000
PubMed
Summary

Efficient gene targeting in Saccharomyces cerevisiae requires sufficient homology. Internal homologous sequences, like hisG, significantly reduce integration efficiency when targeting constructs have limited homology, impacting methods like PCR-mediated gene targeting.

More Related Videos

Targeted in Situ Mutagenesis of Histone Genes in Budding Yeast
08:48

Targeted in Situ Mutagenesis of Histone Genes in Budding Yeast

Published on: January 26, 2017

Removal of an Internal Translational Start Site from mRNA While Retaining Expression of the Full-Length Protein
05:48

Removal of an Internal Translational Start Site from mRNA While Retaining Expression of the Full-Length Protein

Published on: March 16, 2022

Related Experiment Videos

Last Updated: Jul 24, 2026

Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
09:52

Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii

Published on: July 12, 2013

Targeted in Situ Mutagenesis of Histone Genes in Budding Yeast
08:48

Targeted in Situ Mutagenesis of Histone Genes in Budding Yeast

Published on: January 26, 2017

Removal of an Internal Translational Start Site from mRNA While Retaining Expression of the Full-Length Protein
05:48

Removal of an Internal Translational Start Site from mRNA While Retaining Expression of the Full-Length Protein

Published on: March 16, 2022

Area of Science:

  • Molecular Biology
  • Yeast Genetics

Background:

  • Homologous recombination is a key mechanism for gene targeting in Saccharomyces cerevisiae.
  • The hisG cassette is frequently used for gene disruptions, allowing marker recovery but leaving residual sequences.

Purpose of the Study:

  • To investigate the impact of endogenous homologous sequences on gene targeting efficiency.
  • To determine the effect of limited homology in targeting constructs when competing with internal homologous sequences.

Main Methods:

  • Utilizing an integration construct with limited homology at one end for gene targeting in yeast.
  • Comparing integration efficiency with varying numbers of endogenous hisG sequences (0, 1, or 2 copies).
  • Assessing the effect of enlarged homology at the disruption construct.

Main Results:

  • A single genomic hisG copy reduced target integration from 44% to 4.5%.
  • Two genomic hisG copies decreased integration efficiency to below 1%.
  • Increasing homology at the disruption construct abolished the inhibitory effect of endogenous hisG.

Conclusions:

  • Competition between endogenous hisG sequences and targeting constructs with limited homology significantly impairs gene targeting efficiency.
  • Methods relying on limited homology, such as PCR-mediated gene targeting, are less effective in the presence of substantial internal homologous sequences.