Gene Targeting & Transgenic Facility at UCHC

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- Contributed by Deborah Cassarino

 

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DNA Pronuclear Microinjection to Mutant Mouse Embryos

Investigators who have transgene constructs ready should complete a DNA pronuclear microinjection request form.

The microinjection will be scheduled according to the PI's breeding plan.

PI will need to set up enough breeding pairs to supply the facility with 15 KO females of appropriate age (3 weeks old) and separately housed fertile breeding age (8 weeks old) KO males. The GTTF staff will supply hormones for I.P. injection. The PI will arrange to have their staff perform hormone injections when scheduled and set up the breeding. GTTF staff will harvest oviducts the following day for microinjection.
To set up enough breeding pairs, you will need to know the production index of your strain of mice. The production index (PI) is the number of pups per week per female. First you will set up enough breeding pairs to get 15 eight week old males.
#Female X PI = 16 (8 pups will be male) This will give you the number of females you need to set up to get 8 male pups. You will need these to be 8 weeks old for breeding.
Then set up #Female X PI = 30 (15 will be females).

The fee for pronuclear microinjection into knock out (KO) embryos can be found here. We guarantee 3 transgenic mice or 50 pups.

If you are unable to have enough females available, the GTTF can do microinjection on a ‘daily’ basis. The fee can be found here. There will be no guarantee for transgenic mice.

At least 60 ug of circular construct DNA is needed. Please use Endofree plasmid kit or CsCl-Ethidium Bromide gradients method to prepare your plasmid DNA.