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 Experiment Videos

Simple and highly efficient BAC recombineering using galK selection.

Søren Warming1, Nina Costantino, Donald L Court

  • 1Mouse Cancer Genetics Program, National Cancer Institute Frederick, MD 21702-1201, USA.

Nucleic Acids Research
|February 26, 2005
PubMed
Summary
This summary is machine-generated.

Related Concept Videos

You might also read

Related Articles

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

Sort by
Same author

Enhancement of RecET-mediated in vivo linear DNA assembly by a xonA mutation.

PloS one·2026
Same author

A λ Phage Platform for Successful Therapeutic Display of Protein Antigens.

bioRxiv : the preprint server for biology·2026
Same author

The common murine retroviral integration site activating Hhex marks a distal regulatory enhancer co-opted in human early T-cell precursor leukemia.

The Journal of biological chemistry·2025
Same author

Correction: Recombineering: Genetic Engineering in Escherichia coli Using Homologous Recombination.

Current protocols·2024
Same author

Novel mechanisms of MITF regulation identified in a mouse suppressor screen.

EMBO reports·2024
Same author

Stability and gene strand bias of lambda prophages and chromosome organization in <i>Escherichia coli</i>.

mBio·2024
Same journal

Correction to 'scSuperAnnotator: A platform for benchmarking comparison and visualizing automated cellular annotation methods for scRNA-seq data'.

Nucleic acids research·2026
Same journal

Correction to 'Differentiable partition function calculation for RNA'.

Nucleic acids research·2026
Same journal

Deployment of non-canonical splicing in tunicate genomes is mediated by divergent U2AF function and changing m6A modification in U1 and U6 snRNA.

Nucleic acids research·2026
Same journal

Bacillus subtilis DnaB forms multiple protein-protein interactions essential for DNA replication initiation.

Nucleic acids research·2026
Same journal

Multiple forms of protein-protein and DNA binding are exhibited by BrxC from the BREX phage restriction system.

Nucleic acids research·2026
Same journal

Biosynthesis of glycosylated 5-hydroxycytosine in the DNA of diverse viruses.

Nucleic acids research·2026
See all related articles

New recombineering strains enable bacterial artificial chromosome (BAC) modification using efficient galK selection. This method allows precise DNA changes without unwanted markers, facilitating genetic studies.

Area of Science:

  • Molecular Biology
  • Genetics

Background:

  • Recombineering utilizes lambda Red-mediated homologous recombination for DNA modification in Escherichia coli.
  • Existing methods often require restriction enzymes and ligases, or introduce unwanted selectable markers.

Purpose of the Study:

  • To develop novel recombineering strains for efficient bacterial artificial chromosome (BAC) modification.
  • To implement a galK positive/negative selection system for precise DNA engineering without residual markers.

Main Methods:

  • Construction of three new recombineering strains (SW102, SW105, SW106) with a deleted galK gene and a defective lambda prophage.
  • Utilizing galK positive/negative selection for targeted DNA modifications.
  • Incorporating inducible Cre or Flp recombinase genes in strains SW105 and SW106.

Related Experiment Videos

Main Results:

  • Demonstrated the efficiency of galK selection for both positive and negative selection of DNA modifications.
  • Showcased the ability to introduce point mutations, deletions, and loxP sites into BAC DNA.
  • Achieved significantly reduced background compared to other negative selection schemes.

Conclusions:

  • The new recombineering strains and galK selection offer a powerful and efficient tool for BAC engineering.
  • This system facilitates functional studies of mutations, regulatory elements, and the construction of conditional vectors.