CreER-LoxP System-Based Target Gene Inactivation: A Tamoxifen-Inducible Cre Recombinase System for Target Gene Knockout in a Mouse Model

Samenvatting

Source: Hubner, E. K., et al. Constitutive and Inducible Systems for Genetic In Vivo Modification of Mouse Hepatocytes Using Hydrodynamic Tail Vein Injection. J. Vis. Exp. (2018)

This video describes a procedure for target gene knockout using a tamoxifen-inducible Cre-recombinase system following intraperitoneal injection of tamoxifen in a mouse model.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Induction of Transfected CreER with Tamoxifen

CAUTION: Tamoxifen is harmful, may be cancerous or damage fertility. Please refer to the safety data sheet.

1. Plan intraperitoneal tamoxifen injections on three consecutive days.
2. On day 1, dissolve 10 mg of tamoxifen in 40 µL ethanol. Incubate at 55 °C for 10 min. Vortex several times until the tamoxifen has dissolved.
3. Add 960 µL corn oil. Incubate for 5 mins at 55 °C. Vortex several times to get a clear solution.
4. Prepare the solution in a 1-mL insulin syringe with a 27 G needle.
5. Scruff the mouse by grabbing the neck of the mouse carefully with the thumb and the second finger, fixing the tail between the base of the hand and the fourth and fifth finger.
6. Inject 0.1 mL (= 1 mg of tamoxifen) of the solution intraperitoneally into the left lower quadrant of the abdomen.
7. Repeat the injections on days two and three.

Materialen

Lijst van materialen gebruikt in dit artikel

Naam	Bedrijf	Catalogusnummer	Opmerkingen
Injekt Syringe F 1 ml	Braun	9166017V	For intraperitoneal injection
Tamoxifen	Sigma-Aldrich	T5648-1G	For CreER activation
Corn oil	Sigma-Aldrich	C8267-500ML	Carrier for tamoxifen injections