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Updated: Jul 6, 2026

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

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

Published on: March 16, 2012

Site-specific chromosomal integration mediated by phiC31 integrase.

Annahita Keravala¹, Michele P Calos

¹Department of Genetics, Stanford University School of Medicine, Stanford, CA.

Methods in Molecular Biology (Clifton, N.J.)

|March 29, 2008

Summary

The phiC31 integrase system enables precise, targeted gene integration into mammalian cells. This method uses antibiotic selection and PCR to ensure accurate, repeatable gene placement, minimizing position effects.

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Area of Science:

Molecular Biology
Genetics
Biotechnology

Background:

The phiC31 integrase is a bacteriophage-derived site-specific recombinase utilized in mammalian systems.
It mediates unidirectional recombination between specific attB and attP sites.

Purpose of the Study:

To develop a method for precise, targeted integration of plasmids into mammalian chromosomes using phiC31 integrase.
To minimize gene expression variability caused by random integration events.

Main Methods:

Preintegration of an attP site into a mammalian chromosome.
Introduction of a plasmid containing an attB site for recombination with the chromosomal attP site.
Utilizing antibiotic selection and polymerase chain reaction (PCR) for verification.

Main Results:

Achieved precise and unidirectional integration of the plasmid into the targeted chromosomal attP site.
Successfully isolated cell lines with desired integration using antibiotic selection.
Verified correct chromosomal placement via PCR assay.

Conclusions:

The phiC31 integrase system provides a robust method for targeted gene integration in mammalian cells.
This system facilitates reproducible gene insertion at a specific chromosomal locus, reducing position effects.
The described protocol offers a simple approach for obtaining repeated integration at the same site.