Recombinant DNA Experiments
Because pronuclear injections and ES cell electroporations utilize unique DNA sequences that recombine into the mouse genome, the principal investigator needs to provide the GEM Facility with an approval number from the Institutional Biosafety Committee (IBC) prior to beginning these experiments.
Prior to the production of any transgenic or chimeric mice, and to performing cryopreservation, all investigators are required to obtain approval for animal research from the MD Anderson Veterinary Medicine Surgery Division by completing the MD Anderson Animal Care and Use Form (ACUF). The GEM Facility cannot initiate any experiments without a valid animal use protocol number.
All information related to the experiment will be held fully confidential by the GEM Facility. No knowledge related to the experiment will be shared unless otherwise instructed in writing by the principal investigator.
Submission of a Service Request Form with the appropriate Cost Center to charge is required prior to the initiation of service by the GEM Facility. This form will be submitted for billing at the time service is performed.
The MD Anderson Genetically Engineered Mouse Facility is supported, in part, by a Cancer Center Support Grant from the National Cancer Institute. Therefore, all publications resulting from the services provided by the GEM Facility must cite the facility in the acknowledgement section. A copy of all publications resulting from animals created by the GEM Facility should be submitted to Jan Parker-Thornburg.
Service will be scheduled upon the receipt of all pertinent forms and documentation. All services will be provided according to the order in which requests were received.
Specific Policies for Experiments
ES cells provided to the facility will be injected after visual inspection of their morphology. Cells that are in poor condition will not be injected, and investigators may be charged an additional fee for blastocysts collected. To avoid unnecessary animal use, we request a week's notice for cancellation of an injection date.
Gene-Targeting in ES Cells
The investigator is responsible for demonstrating a reliable scheme for identifying targeted ES cell clones by Southern analysis with 5' and 3' probes by providing results from pilot experiments for this purpose. PCR analysis may be suitable for analysis if appropriate control experiments have been done. (Call Jan Parker-Thornburg for further information on certifying PCR analysis.) A convincing strategy for identifying mutant alleles must be demonstrated prior to the initiation of a gene-targeting experiment.
Our facility has produced thousands of founder lines by pronuclear injection. While DNA can generally be shown to incorporate into the genome, the gene may or may not exhibit expression. It is not always possible to predict whether the transgene will be expressed and the effects of its expression in the mouse. For this reason, the GEM Facility does not guarantee transgene expression. The investigator is responsible for demonstrating a reliable scheme for identifying detection of the transgene at a single copy level by providing results from pilot experiments for this purpose. The GEMF will provide protocols for single-copy detection upon request.
While cryopreservation is a valuable tool for conserving mouse resources, certain mouse strains are not amenable to the rigors of the technique. For that reason, it is imperative to know the genetic background of the strain to be cryopreserved. The GEM Facility will thaw one straw of frozen embryos and culture overnight to the blastocyst stage to insure that embryo freezing is feasible for that particular mouse line. For sperm cryopreservation, the investigator will be given a schedule for pre-mating the males to encourage new sperm production.