Mission of the Murine Models Core and services provided.

The Murine Models Core provides assistance to DDRCC investigators for use of the mouse as an experimental model. The Director of the Core is Jeff Miner, and the co-Director is Peter Crawford. This Core provides project advice and technical services that include production of transgenic and chimeric mice, derivation of speed congenics, and assisted reproduction services for maintaining pedigrees via the Mouse Genetics Core, as well as gnotobiotic mouse provision, colonization, and rederivation. The Murine Models Core provides timely access to these services at a reasonable cost to DDRCC investigators, as well as consultations with investigators who need information regarding project design or technical implementation of these models.

SERVICE REQUESTS

 

Requests for Mouse Genetics Core services should be made on the MGC services request form. The appropriate MGC form should also be filled out. Submit both forms to Jeff Miner (Box 8126). Requests for services not listed are welcomed, as are queries concerning project design or the details for any service.
Requests for gnotobiotic services should be made on the gnotobiotic services request form and submitted to both Jeff Miner (Box 8126) and Peter Crawford (Box 8510). For colonization and rederivation services, the project description requested below must also be submitted.

 

SPECIFIC SERVICES OFFERED

 

The DDRCC Murine Models Core offers a variety of services to assist investigators with production and use of murine models for gastrointestinal research. Investigators are encouraged to contact Jeff Miner for information/advice about any service, including potential services not listed. The following services are provided:

 
 

TRANSGENIC MOUSE PRODUCTION

Investigators construct a transgene or BAC and purify the DNA fragment to be microinjected using an approved protocol. The core will inject the construct into C57BL/6J, FVB/N, (C57BL/6JxCBA)F2 mice, or other strains as requested. The DDRCC investigator cost is $1000.00.

 
 

CHIMERIC MOUSE PRODUCTION

For production of mice with targeted mutations (knockout/knockin and chimeric-transgenic mice), the Murine Models Core will inject embryonic stem cells into blastocysts for production of chimeric animals. Investigators produce targeting constructs, and ES cell transfection can be performed by the investigator or by the Siteman Cancer Center Mouse Embryonic Stem Cell Core. Cells for injection prepared at the Mouse Embryonic Stem Cell Core are transferred directly to the DDRCC Murine Models Core for chimeric mouse production after the investigator identifies the targeted ES cell clones. This service provides enough injections to obtain the desired chimeras. The DDRCC investigator cost is $1,000.00 per mutation.

 

PROVISION OF RNA FROM GNOTOBIOTIC MICE

RNA has been prepared from gastrointestinal tissues of matched gnotobiotic and control C57BL/6J mice at 6-12 weeks of age. The DDRCC investigator cost is $50.00 for each set of RNA samples.

 

PROVISION OF GNOTOBIOTIC MICE

Matched pairs of C57BL/6, FVB/N, or C57BL/6 Rag1-/- gnotobiotic/conventional animals are available for DDRCC investigators. A breeding cycle (four weeks until birth plus the age of the mice desired) may be necessary before these animals are available. Cost to DDRCC investigators is $10.00 per pair.

 

GNOTOBIOTIC COLONIZATION AND RE-DERIVATION

Germ free mice can be colonized with normal human or mouse gut microbiota, or with pathogenic bacteria for defined periods, usually 4-8 weeks. DDRCC investigator cost is $94.00 per colonization (a ÔcolonizationÕ is defined as a single isolator of animals, which can harbor up to 5 cages) plus gnotobiotic housing costs ($1.09/cage/day to the investigator) and germ free mouse costs (see above). Moreover, conventionally raised mouse strains can be re-derived as germ free: DDRCC investigator cost is $1092 per pedigree re-derived. Colonization and rederivation services require a substantial commitment of Core resources. Requests for gnotobiotic colonization and rederivation services will be overseen by a review committee (Jeffrey Gordon, Phillip Tarr, William Stenson, Peter Crawford). DDRCC members interested in these services should submit a written proposal along with the gnotobiotic services request form within 4 weeks of one of three call dates: April 15, August 15, and December 15. The proposal should be less than two pages and include a statement of the hypothesis being tested, rationale, definition of experimental design, and precise details for the number of experimental and control animals needed and duration of colonization. Each proposal should be confined to a specific experimental aim. If these experiments are a component of or the precursor to additional studies and a larger project, the overall goals of the study and additional anticipated experiments should be briefly summarized in the proposal. Investigators are strongly encouraged to contact Peter Crawford before submitting a proposal.

 

CRYOPRESRVATION

Murine embryos are vitrified and preserved indefinitely in liquid nitrogen. This technique allows strains of mice to be preserved for future use without the labor and expense of continuous mating, and preserves a strain from risk of pathogen infection. This service includes cryopreservation of up to 300 embryos and includes a test thaw and implantation. The cost to DDRCC investigators is $300.00 for embryo freezing and $25.00/year/strain for preservation.

 

ASSISTED REPRODUCTION

In vitro fertilization is offered to assist propagation of murine pedigrees where a male mouse is too old or is otherwise unable to produce offspring through natural mating. Ovarian transplant and/or superovulation of female animals to assist in propagation of murine pedigrees with breeding defects can be performed. The cost to DDRCC investigators is $150.00 for IVF and $20.00 for each ovarian transplant donor.

 

SPEED CONGENICS

Targeted mutations in one genetic background (for example, 129SV) frequently need to be transferred to another background more suitable for physiological analysis (for example, C57BL/6J). This process takes at least ten generations through natural matings, but strain-specific polymorphic markers can be utilized to speed this process to no more than five generations. Offspring from each round of mating that have the greatest contribution of chromosomes from the desired background are selected for mating to produce the next generation. This selected breeding process is termed "speed congenics." The core uses single-nucleotide polymorphisms to distinguish between strains, and conversion between most inbred lines can be accommodated. The five generations take approximately fourteen months to complete moving a mutation from one inbred strain to another. The investigator provides several males and at the end of the conversion will be returned at least one male animal in the recipient background. The cost to DDRCC investigators for this service is $2,000.00 per conversion or 20% of the actual cost, whichever is less.

PROTOCOLS

  TRANSGENE PREPARATION PROTOCOL
UNIVERSAL GENOTYPING PROTOCOL

OUTREACH

  The DDRCC Murine Models Core sponsors a gnotobiotic model interest group which meets periodically to discuss ongoing experiments, explore new technological developments, and offer feedback for proposed studies. Contact Peter Crawford for inclusion in the mailing list and meeting information.