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Updated: Jun 15, 2026

Generation of Enterobacter sp. YSU Auxotrophs Using Transposon Mutagenesis

Generation of Enterobacter sp. YSU Auxotrophs Using Transposon Mutagenesis

Published on: October 31, 2014

A practical random mutagenesis system for probiotic Lactobacillus casei using Tn5 transposition complexes.

M Ito¹, Y-G Kim¹, H Tsuji²

¹Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.

Journal of Applied Microbiology

|March 6, 2010

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A new Tn5 transposome system efficiently creates random genetic mutations in Lactobacillus casei (L. casei). This tool enables the construction of a large mutant library, aiding research into L. casei

Area of Science:

Microbiology
Genetics
Probiotics

Background:

The genome of probiotic Lactobacillus casei (L. casei) is sequenced, but its beneficial mechanisms remain unclear due to a lack of efficient mutagenesis systems.
Understanding L. casei's genetic basis is crucial for harnessing its full probiotic potential.

Purpose of the Study:

To establish a practical random mutagenesis system for L. casei using Tn5 transposome complexes.
To create a comprehensive Tn5 insertion library for L. casei.

Main Methods:

Optimized transformation conditions using plasmid pUCYIT356-1-Not2.
Developed a transposition reaction protocol using the Tn5 transposome system for L. casei ATCC 27139.
Constructed a library of 9408 Tn5 insertion mutants and analyzed auxotrophic mutants via inverse PCR to determine insertion sites.

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Creation of a Dense Transposon Insertion Library Using Bacterial Conjugation in Enterobacterial Strains Such As Escherichia Coli or Shigella flexneri

Published on: September 23, 2017

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Last Updated: Jun 15, 2026

Generation of Enterobacter sp. YSU Auxotrophs Using Transposon Mutagenesis

Generation of Enterobacter sp. YSU Auxotrophs Using Transposon Mutagenesis

Published on: October 31, 2014

Creation of a Dense Transposon Insertion Library Using Bacterial Conjugation in Enterobacterial Strains Such As Escherichia Coli or Shigella flexneri

Creation of a Dense Transposon Insertion Library Using Bacterial Conjugation in Enterobacterial Strains Such As Escherichia Coli or Shigella flexneri

Published on: September 23, 2017

Main Results:

Successfully established and optimized a Tn5 transposome-based mutagenesis system for L. casei.
Generated a large-scale Tn5 insertion library comprising 9408 mutants.
Confirmed random Tn5 insertion across the L. casei genome, demonstrated by the isolation and analysis of six auxotrophic mutants.

Conclusions:

The Tn5 transposome system is an efficient method for generating transposon insertion mutants in L. casei.
The developed system facilitates the construction of a valuable L. casei Tn5 insertion library.
This mutagenesis system will advance research into the beneficial effects of L. casei for human health.