Related Experiment Video

Updated: Jun 6, 2026

Site-specific Bacterial Chromosome Engineering: ΦC31 Integrase Mediated Cassette Exchange (IMCE)
08:21

Site-specific Bacterial Chromosome Engineering: ΦC31 Integrase Mediated Cassette Exchange (IMCE)

Published on: March 16, 2012

I-SceI-mediated plasmid deletion and intra-molecular recombination in Spiroplasma citri

Marc Breton1, Sybille Duret, Laure Béven

  • 1INRA, Génomique Diversité et Pouvoir Pathogéne, Villenave d'Ornon, France.

Journal of Microbiological Methods
|December 7, 2010
PubMed

Insights

Researchers utilized meganuclease I-SceI for plasmid deletion in Spiroplasma citri. This method enabled targeted removal of plasmids, offering a new tool for genetic manipulation in spiroplasmas.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • Spiroplasma citri (S. citri) harbors six plasmids (pSci1-6) potentially involved in transmission by leafhopper vectors.
  • Understanding and manipulating these plasmids is crucial for studying S. citri biology and vector interactions.

Purpose of the Study:

  • To investigate the use of meganuclease I-SceI for targeted plasmid deletion in S. citri.
  • To establish I-SceI as a counter-selection tool for genetic engineering in mollicutes.

Main Methods:

  • Introduction of engineered plasmids (pSci1NT-I, pSci6PT-I) containing I-SceI recognition sites and a tetM marker into S. citri strains.
  • Transformation with an oriC plasmid expressing the I-SceI gene under the spiralin promoter.
  • Analysis of plasmid loss and deletion events mediated by I-SceI expression.

Main Results:

  • I-SceI expression led to the rapid loss of a target plasmid (pSciNT-I) in S. citri strain 44, demonstrating its utility as a counter-selection marker.
  • In S. citri strain GII3, I-SceI expression induced deletion of plasmid fragments containing the I-SceI site and tetM marker.
  • Deletions were confirmed to occur via intramolecular recombination between homologous sequences.

Conclusions:

  • Meganuclease I-SceI can be effectively used for plasmid deletion in S. citri.
  • This study reports the first instance of I-SceI-mediated intramolecular recombination in mollicutes.
  • The findings provide a novel method for genetic manipulation of S. citri plasmids.

More Related Videos

Subcloning Plus Insertion (SPI) - A Novel Recombineering Method for the Rapid Construction of Gene Targeting Vectors09:02

Subcloning Plus Insertion (SPI) - A Novel Recombineering Method for the Rapid Construction of Gene Targeting Vectors

Published on: January 8, 2015

Application of CRISPR Interference (CRISPRi) for Gene Silencing in Pathogenic Species of Leptospira14:49

Application of CRISPR Interference (CRISPRi) for Gene Silencing in Pathogenic Species of Leptospira

Published on: August 14, 2021

Related Concept Videos

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...
Bacterial Phylum Tenericutes01:24

Bacterial Phylum Tenericutes

The phylum Tenericutes, which includes the single class Mollicutes, comprises bacteria that lack cell walls. The term "Mollicutes" derives from the Latin word mollis, meaning "soft." These organisms are among the smallest known and are commonly referred to as mycoplasmas due to the prominence of the genus Mycoplasma, which includes well-known human pathogens. Despite their inability to stain gram-positively (a result of their lack of cell walls), mycoplasmas are phylogenetically related to the...