Gene Targeting Lab

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Expert:

Behrouz Asgari (MSc)


Gene Targeting Laboratory: Transgenic Mouse Production and Services

The establishment of transgenic mouse production technology is indispensable in major research centers, as these models serve as powerful tools for a wide range of studies, including developmental biology, regenerative medicine, cell therapy, and other biological fields. Accordingly, the Gene Targeting Laboratory was established at the Royan Institute to develop transgenic mouse models.

The foundation of transgenic mouse technology was laid by two key scientific advancements:

  1. The ability to manipulate early-stage mouse embryos in vitro.

  2. Advances in gene transfer techniques into mouse embryonic stem cells (ESCs).

Using microinjection of transgenic ESCs into mouse embryos, the laboratory successfully produced Iran’s first transgenic mouse ("Green Mouse"). Subsequent achievements include the Pdx1-EGFP mouse (with green fluorescent insulin-secreting cells) and the country’s first CRISPR/Cas9-mediated Knock-In mouse.

Available Transgenic Mouse Models

  1. Tg(CAG-EGFP) ("Green Mouse")

    • All cells express EGFP (enhanced green fluorescent protein).

    • Puromycin-resistant, enabling antibiotic selection.

  2. Tg(Pdx1-EGFP)

    • EGFP expression restricted to pancreatic β-cells (under the Pdx1 promoter).

    • Fluorescent labeling of Langerhans islet cells.

  3. Zfp521 Knock-In (CRISPR/Cas9)

    • Inducible Zfp521 expression (drives neural differentiation).

    • Integrated into the ROSA26 locus for ubiquitous activation.

    • Enables somatic-to-neural stem cell reprogramming.

  4. mT/mG Knock-In (CRISPR/Cas9)

    • Dual-fluorescence reporter:

      • Ubiquitous mRFP (red).

      • Cre-dependent mGFP (green) (upon mRFP excision).

    • Ideal for lineage tracing and cell fate mapping.

Core Services

1. Microinjection & Genetic Engineering

  • Pronuclear injection of foreign DNA into mouse zygotes.

  • Zebrafish zygote microinjection (transgenesis).

  • ESC injection into mouse blastocysts (chimeric mouse production).

2. Embryo Manipulation

  • Aggregation: Embryonic stem cell-morula fusion.

  • Electroporation: Foreign DNA delivery into cells.

  • Embryo transfer (uterus/oviduct of pseudopregnant mice).

3. Embryo & Strain Preservation

  • Preimplantation embryo isolation (all developmental stages).

  • Genotyping of transgenic mice.

  • Cryopreservation of embryos/sperm (for archiving or import/export).

Research Impact

The laboratory supports dozens of projects (both within Royan Institute and externally) by providing:

  • Custom transgenic/KO mouse models.

  • CRISPR/Cas9-mediated genome editing.

  • Disease modeling (cancer, diabetes, neurogenesis).

  • Cell lineage tracking tools (e.g., mT/mG).

This facility bridges genetic engineering with in vivo biomedical research, enabling studies that were previously unfeasible in Iran.