RRC-TPS: User Policies

From RRC
Jump to: navigation, search

TPS Facility User Policies

Please visit the main web page of the Transgenic Production Service for an overview of our services.

For additional service-specific instructions and related policies refer to the appropriate service page: DNA Microinjection, ES Cell Microinjection, Gene Targeting in ES Cells, Cryopreservation.

Transgenic mouse, embryonic stem (ES) cell, and cryogenic services are performed on a first-come, first-served basis. A valid RRC account, to which all Production Fees are charged, is required before any service is initiated. Fees for animal purchases and per diem are charged separately to the investigator's BRL account number, which must be submitted to the TPS when service is requested. Estimated total fees for most services are posted. For special-needs projects, cost estimates will be provided to investigators for approval after consultation with TPS staff. In addition, BRL may request mouse pathogen screening for animals moved from the TPS to certain housing areas on campus. These tests are also charged directly to the BRL account. For outside users, BRL fees will be charged to the RRC account on a re-charge basis. All projects for which transgenic or chimeric mice will be generated, or for which any mice will be transferred to an investigator, require an approved ACC Animal Protocol describing the relevant transgene, gene modification, or mouse strain. The Approval number must be submitted to the TPS Director at the time service is requested. Outside investigators must submit their institution's Approval letter.

DNA fragments (5-10 ug) submitted for microinjection into embryos to produce transgenic founders should be highly purified and at a concentration of about 50-100 ng per microliter. Refer to our protocol for preparing DNA for microinjection. Before mice are ordered and a project scheduled the TPS will confirm DNA concentration, size and integrity by gel electrophoresis. While the rate of transgenesis varies between DNAs, the TPS averages five founders per construct. If two founders are not generated during the first round of injections, additional injections may be performed if evidence is provided to show that the screening assay is working. In this case the RRC Production Fee will be waived but the investigator will pay for the required mice and per diem. A freshly prepared DNA sample may be requested. Note that the TPS reserves the right to abort any project during the first round of injections if the DNA is determined to be unsuitable (i.e., contains particulates) for microinjection. If a project is halted BRL animal fees will still be charged as will 60% of the Production Fee.

ES cells submitted for microinjection to produce chimeras must have tested negative for mycoplasma (testing available at the DNAS facility) and have been screened for a panel of mouse pathogens. Documentation of these tests must be submitted ahead of time. Contact TPS for details. On each day of microinjection the investigator will provide ES cells in suspension at a scheduled time. Cancellations will result in a charge of 100% of animal fees and 60% of the Production Fee. For an additional fee, cells may be submitted as frozen stocks to be expanded and prepared for microinjection by the TPS. In this case, if cells have not yet been tested for MPV the facility will arrange for testing (fees will apply). The TPS also offers gene targeting services, and maintains germline-competent ES cell lines for these projects. Because the generation of chimeras and subsequent germline transmission is highly dependent on several factors, which include culture conditions, chromosome complement and acquisition of secondary mutations during culture, we cannot guarantee success with all cells.

For gene targeting experiments the investigator must provide electroporation targeting DNA (150 ug - 300 ug, linearized) at a minimum concentration of 1 ug per microliter (protocol). Evidence of a sensitive screening assay for analysis of homologous recombination (HR) must also be submitted in advance of electroporation. Prior consultation with TPS staff is highly recommended to assure that targeting vectors are isogenic with the germline-competent ES cells available in the facility. Investigators are responsible for screening drug-selected ES clones (at least 288) for HR, which must be performed within two months. Potentially targeted clones identified in this initial screen will be expanded and DNA supplied to the investigator to confirm HR. As HR is highly dependent on the locus being modified, we cannot guarantee that targeting will occur with a given construct.

Fees are posted in the Facility Fees Index.

All specialized media and reagents not normally stocked by the TPS facility will be charged to the investigator.

We reserve the right to refuse projects that involve use of reagents, techniques or animals that we deem hazardous.

Please acknowledge the Transgenic Production Service if you use mice produced in our facility in a publication.

Xenogen IVIS Spectrum User Policies

IVIS User Policies

Personal tools