ES Cell Microinjection

1. The investigator must open an RRC account (the RRC Application may be downloaded from the RRC website and the completed form faxed to RRC) and a BRL account (contact the BRL Office).
   
   
2. The investigator must submit a UIC Animal Use Protocol for approval by the Animal Care Committee (ACC) before initiation of this project. When preparing your Protocol please refer to the existing TPS ACC Protocol (contact Dr. Franks) for generating chimeric mice. Any plans for experimental usage of mice subsequent to the generation of chimeras by our Facility must be described in your Protocol application and approved by the committee.
   
   
3. The investigator must complete the Application for ES Cell Microinjection (application may be downloaded from TPS website).
   
   
4. When the completed Application form is submitted to the TPS, the investigator and Director will discuss the necessary preparations and arrange a time for performing the microinjections. The investigator will also be asked at this time (if not earlier) to sign a letter giving the TPS permission to order mice under the investigator's BRL account. Mice will be ordered to arrive 1/2- to 1-week ahead of time. The TPS will take responsibility for keeping an accurate daily account of the number of mice being housed under the investigator's BRL account.
   
   
5. Generally, embryos will be microinjected with ES cells carrying a given targeted mutation and transplanted into pseudopregnant females over a 5-day period. A single targeted clone may be microinjected but we recommend that you provide 2-3 clones, as clones may vary with respect to their potential for germline transmission. In addition, the generation of at least 2 independent lines (2 distinct clones) of mice with a given targeted mutation provides a control for your studies. Approximately 120 injected embryos are transplanted into 10 foster (surrogate) mothers, although variability with regard to these numbers should be expected (note, that occassionally due to unforseen problems additional mice may have to be ordered and injections carried out over a longer period of time)
   
   
6. The investigator will be notified when the microinjections have been completed and when the potential litters are due (17-19 days after microinjection).
   
   
7. At weaning (3 weeks) chimeric mice will be transferred out of the TPS to the care of the investigator. Prior to this time please make the appropriate arrangements for housing your mice. Please note: The number of chimeras produced is dependent on the quality of, and the chimeric rate for, the submitted ES cells. We cannot guarantee success with every ES clone.
   
   
8. At approximately 6 weeks of age chimeras are ready for mating with C57BL/6 mice for determination of germline transmission.