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The Tulane Transgenic Mouse Facility |
Transgenic Mouse Production by Pronuclear Microinjection
Pronuclear injection is the most common method used to create transgenic mice. This procedure involves collecting fertilized eggs at the single cell stage. For a brief window of time the pronuclei containing the genetic material from the sperm head and the egg are visible within the protoplasm. At this stage, a linearized DNA construct is injected into one of the pronuclei. The injected eggs are then transferred into the oviducts of pseudopregnant foster mice. Generally 10 to 20% of the pups born to the foster mothers have integrated the injected DNA into their genomes, thus becoming transgenic. Each pup is a unique founder mouse, as the DNA integrates randomly into the genome.
The transgenic mouse technique is the most commonly used method to study "gain-of-function" of a gene in mouse. The idea is to force expression of a gene and study its consequences. This technique addresses whether ectopic or over-expression of a gene is sufficient to cause anomalies. The facility can generate transgenic mice for your research needs.
Investigators must supply the following items
- Completed Request for Service form
- Digested DNA OR a gel slice containing the linearized fragment and an image of a gel verifying the constructÕs size and complete digestion (The investigator is responsible for getting approval from the Institutional Biosafety Committee for using recombinant DNA)
- Current protocol number for the transgenic pups
- IT form for facility charges (Tulane only)
General timeline
- DNA delivered
- DNA purified for microinjection
- Eggs injected for 3 sessions (investigator notified)
- Foster mice give birth (investigator notified)
- Tail tips given to investigator at 15-16 days
- Pups are weaned at 21 days and transgenic pups are released to the investigator
- The time from notification of beginning injections sessions to delivery of tail tips normally takes 5-8 weeks, with 3 sets of tails expected.
Special notes
The facility guarantees the production of 3 transgenic pups per construct, however there is no guarantee that the pups produced will have germline transmission or expression of the transgene.
The facility routinely uses the hybrid strain B6C3F1 from J as the background strain. Investigators who need to use a different strain will separately purchase the male and female mice required for the procedure and pay the per diem charges for these mice.
If the investigator has not contacted the facility with the results of the genotype by weaning (21 days), all the pups will be transferred to the investigator's protocol number and the investigator must pay the per diem charges for the pups.
The investigator must inform the facility if the transgene is expected to produce severe defects or lethality, so that the pups can be properly monitored.
If your facility requires specific health tests before they will accept the transgenic pups, the investigator will pay for the cost of testing, including any shipping charges.
The current charge for pronuclear injection is $1200 for Tulane investigators, $1500 for outside investigators.
Last updated May, 2008