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

Development of a lactococcal integration vector by using IS981 and a temperature-sensitive lactococcal replication

K M Polzin1, L L McKay

  • 1Department of Microbiology, University of Minnesota, Minneapolis 55455.

Applied and Environmental Microbiology
|February 1, 1992
PubMed
Summary

A novel integration vector, pKMP10, facilitates Campbell-like integration into Lactococcus lactis chromosomes. This method enables stable genetic modification of L. lactis strains for research and industrial applications.

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

Scanning Electron Microscopy of Microbial Attachment to Milk Contact Surfaces <sup>1</sup>.

Journal of food protection·2019
Same author

An unusual polymorphic locus useful for tagging Rps1 resistance alleles in soybean.

TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik·2013
Same author

Characterization of the Highly Autolytic Lactococcus lactis subsp. cremoris Strains CO and 2250.

Applied and environmental microbiology·2006
Same author

Influence of calcium and phosphorus, lactose, and salt-to-moisture ratio on Cheddar cheese quality: changes in residual sugars and water-soluble organic acids during ripening.

Journal of dairy science·2006
Same author

Examination of lactococcal bacteriophage c2 DNA replication using two-dimensional agarose gel electrophoresis.

Gene·2001
Same author

Identification of a gene cluster encoding Krebs cycle oxidative enzymes linked to the pyruvate carboxylase gene in Lactococcus lactis ssp. lactis C2.

Journal of dairy science·2000

Area of Science:

  • Microbiology
  • Molecular Biology
  • Bacteriology

Background:

  • Lactococcus lactis is a key bacterium in dairy fermentation.
  • Stable genetic modification of L. lactis is crucial for improving its industrial applications.
  • Existing methods for genetic manipulation can be inefficient or unstable.

Purpose of the Study:

  • To construct and characterize a novel integration vector, pKMP10, for Campbell-like integration into the Lactococcus lactis chromosome.
  • To evaluate the efficiency and stability of pKMP10 integration in different L. lactis strains.
  • To investigate the integration sites of pKMP10 within the L. lactis genome.

Main Methods:

  • Construction of the pKMP10 vector using components from pSK11L, IS981, and pVA891.
  • Temperature-sensitive maintenance of pKMP10 in L. lactis subsp. lactis LM0230.

Related Experiment Videos

  • Selection and isolation of pKMP10 integrants under varying temperature and selection conditions.
  • Southern hybridization to analyze integration sites and structures.

    • Main Results:

    • pKMP10 demonstrated successful Campbell-like integration into the L. lactis LM0230 chromosome, with integrants comprising 8-75% of resistant cells.
    • Integration efficiency varied across different L. lactis strains, with lower frequency in MG1363 and no integration in the Rec-deficient MMS368.
    • Southern hybridization revealed integration into five chromosomal sites, four associated with IS981 sequences and one uncharacterized site.
    • Integrated pKMP10 exhibited significantly higher stability compared to unintegrated plasmid.

    Conclusions:

    • The pKMP10 vector is effective for Campbell-like integration into the L. lactis chromosome, particularly targeting IS981 sequences.
    • The temperature-sensitive nature of the vector facilitates selection of integrants.
    • This tool offers a stable method for genetic modification of L. lactis, advancing its use in biotechnology.