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Pronuclear Injection

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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. Please contact Ann Mullin with questions about pronuclear injection services.

Investigators must supply the following items

  • Completed Service Request form
  • Digested DNA and an image of a gel verifying the construct's size and complete digestion
  • Current protocol number for the transgenic pups
  • IT form for facility charges (Tulane only)

General timeline

  • Purified DNA delivered
  • 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 inbred mouse strain C57BL/6 as  background strains. 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), 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.

Please send an e-mail to Ann Mullin for pricing information. Outside investigators must be at an institution based in Louisiana.

 

 

Tulane University, New Orleans, LA 70118 504-865-5000 website@tulane.edu