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Related Experiment Video

Updated: Jun 18, 2026

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

Published on: March 12, 2018

hESC engineering by integrase-mediated chromosomal targeting.

Ying Liu¹, Uma Lakshmipathy, Ali Ozgenc

¹Invitrogen Corporation, Carlsbad, CA, USA.

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

|November 13, 2009

Summary

This study presents a novel retargeting system for human embryonic stem cells (hESCs) using bacteriophage recombinases. The method enables efficient, long-term transgene expression in pluripotent and differentiated cells.

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Last Updated: Jun 18, 2026

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy

Published on: March 12, 2018

Non-Viral Engineering of Primary Human T Cells via Homology-Mediated End-Joining Targeted Integration of Large DNA Templates

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Published on: May 9, 2025

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

Area of Science:

Stem cell biology
Molecular biology
Gene engineering

Background:

Bacteriophage recombinases offer precise genomic integration in human embryonic stem cells (hESCs).
Efficient long-term transgene expression is crucial for stem cell applications.

Purpose of the Study:

To describe a retargeting system for hESCs using phiC31 integrase and R4 integrase.
To enable complex genetic element assembly and controlled transgene expression.

Main Methods:

Engineered hESC line with a chromosomal target for R4 integrase.
Multisite Gateway technology for complex element assembly.
Retargeting using phiC31 and R4 integrases.

Main Results:

High-efficiency targeting of specific loci in hESCs.
Sustained transgene expression over long-term culture and differentiation.
Appropriate regulation of transgenes in both pluripotent and differentiated cells.

Conclusions:

The described system allows rapid assembly of complex plasmid-based assays.
Controllable insertion and active transgene expression in hESCs.
Facilitates versatile applications in stem cell research and therapy.