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Related Experiment Video

Updated: May 24, 2026

Loss-of-Function Approach in the Embryonic Chick Retina by Using Tol2 Transposon-Mediated Transgenic Expression of Artificial microRNAs

Loss-of-Function Approach in the Embryonic Chick Retina by Using Tol2 Transposon-Mediated Transgenic Expression of Artificial microRNAs

Published on: May 18, 2022

A tightly controlled conditional knockdown system using the Tol2 transposon-mediated technique.

Tokuichi Iguchi¹, Hideshi Yagi, Chen-Chi Wang

¹Division of Cell Biology and Neuroscience, Department of Morphological and Physiological Sciences, Faculty of Medical Sciences, University of Fukui, Fukui, Japan.

|March 20, 2012

Summary

This summary is machine-generated.

This study introduces a novel inducible knockdown system for precise gene silencing in the brain, enabling research in later developmental stages and the adult brain without prior knockdown interference.

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In vivo Application of the REMOTE-control System for the Manipulation of Endogenous Gene Expression

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Published on: March 29, 2019

Area of Science:

Neuroscience
Molecular Biology
Genetics

Background:

In utero electroporation is valuable for studying gene function in cortical development.
Challenges exist in applying knockdown to later brain development due to plasmid dilution and degradation.
Previous methods struggle to isolate knockdown effects from earlier developmental influences.

Purpose of the Study:

To develop a tightly controlled, inducible knockdown system for investigating gene function in later brain development and adult stages.
To overcome limitations of existing methods, such as plasmid dilution and confounding early knockdown effects.

Main Methods:

Developed a novel vector, pT2K-TBI-shRNAmir, utilizing Tol2 transposon-mediated gene transfer and tetracycline-inducible gene expression.
Demonstrated stable transgene maintenance and inducible gene knockdown.
Applied the system to knockdown amyloid precursor protein (APP) in PC12 cells and Dab1 in the developing cerebral cortex.

Main Results:

Successfully achieved sharp reduction of endogenous amyloid precursor protein (APP) expression in PC12 cells.
Induced acute Dab1 insufficiency, leading to impaired radial migration in the developing cerebral cortex.
Confirmed tight control of knockdown with no expression leakage in vivo, as effects were only observed upon induction.

Conclusions:

The developed inducible knockdown system allows for gene function investigation at any stage of brain development, including adult stages.
This method overcomes limitations of plasmid dilution and degradation, enabling stable and long-term gene silencing.
Enables knockdown analyses free from the influence of confounding earlier knockdown effects, providing cleaner experimental results.