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Pronuclear Injection

Foreign DNA, when injected into the pronucleus of a fertilized egg, can integrate into the genome and the resulting mouse will generally carry copies of the transgene in every cell, including germ-cells. The number of transgenes incorporated into the genome can vary both in the number of copies integrated and in the direction of integration.

GEMF Service

A DNA construct, provided by the investigator and purified by the facility, will be injected into fertilized mouse eggs and transferred into the oviduct of foster females. Resulting pups are potentially transgenic and will be transferred to the investigator for analysis.

Cost

$1500/construct for MD Anderson investigators; $4090/construct for external investigators

Time Line for Pronuclear Injection

  • Day 1: Microinject DNA
  • Day 19: Pups born
  • Day 33: Investigator notified about births, male/female numbers. Investigator is responsible for turning in an animal order form for these mice to Veterinary Medicine at this time
  • Day 45: Wean and transfer all pups born from injections to the investigator (upon receipt of signed order from Veterinary Medicine)

To Obtain Service from the GEMF

  • Obtain authorization for animal research from the Office of Research Administration (animal protocol coordinator at Ext. 2-3220). The facility cannot initiate any experiment without an authorized animal research protocol number.
  • Schedule an appointment with Jan Parker-Thornburg (713-745-2654, or e-mail jpthorn@mdanderson.org) to set up an injection date, for protocols on how to prepare the plasmid DNA and for other requisite forms for service
  • Demonstrate the ability to detect a single copy of the gene of interest in the background of genomic DNA. Protocols for determining copy level standards can be found at The University of Michigan's TAMC Transgene copy standards Web site. For protocols on how to incorporate these standards into a PCR or Southern analysis, please go to PCR analysis.
  • Complete and submit the service request form for pronuclear injection, a picture of the DNA prepared for fragment isolation, a picture of a PCR or Southern analysis demonstrating detection down to the single copy level and 15-20 mg of the digested DNA plasmid
    Service request form for MD Anderson investigators (pdf)
    Service request form for external investigators (pdf)
  • Obtain and complete an authorization form with Veterinary Medicine for animal orders

After pronuclear injections and implants have been done and pups have been born, the PI must perform tail analysis and convey the results to the facility. All pups will be transferred to the colony of the investigator.

Read the Policies and Procedures section for important information regarding services rendered by the GEMF.


© 2014 The University of Texas MD Anderson Cancer Center