DNA Microinjection

1. The investigator must open both 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 transgenic founders. Any plans for experimental usage of transgenic mice subsequent to the generation of founders by our Facility must be described in your Protocol application and approved by the committee.
   
   
3. The investigator must complete the Application for DNA Microinjection (application may be downloaded from TPS website). In addition, the investigator must read our protocol for Preparation of DNA for Injection into Mouse Eggs; request from the TPS an aliquot of Injection Buffer, in which the final DNA prep must be dissolved; and prepare the DNA fragment for microinjection.
   
   
4. When the prepared DNA, accompanied by a completed Application form, is submitted to the TPS the investigator will be informed as to when the microinjections will be performed. 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. Embryos will be microinjected with a single DNA construct and transplanted into pseudopregnant females over a 4-day period. Approximately 200 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 (19 days after microinjection). If the investigator desires to follow the pups from birth, please let the TPS know at this time so the appropriate arrangements can be made. We encourage investigator participation to identify phenotypes associated with transgene expression in founders.
   
   
7. The TPS will monitor the pregnancies. The investigator will again be notified when all the recipient mothers have delivered (approximately 60% of recipients carry the microinjected embryos to term; litter size is variable). Tail biopsies will be performed at weaning (~21 days postbirth). When all litters have been weaned the tails will be turned over to the investigator for DNA isolation and transgene analysis. Nontransgenic tails will be included for negative controls. You may wish to refer to our reference for A Simple Prodedure for Isolating Tail DNA for Transgenic Analysis. We strongly recommend that you read our comments on Transgenic Screening and make sure the appropriate positive controls are included in your screening strategy. Note that per diem will be charged during the time it takes the investigator to complete the analysis.
   
   
8. Once the tail DNA analysis is complete the investigator will arrange to meet with a member of the TPS to identify the founders and to transfer them out of the TPS to the care of the investigator. Prior to this time please make the appropriate arrangements for housing your mice.