Transgenic Research Center In the Cancer Center

Mouse rederivation for SIM1 barrier facility

 

Purpose:

All mice that are brought into the SIM1 barrier facility need to be re-derived, except for mice that are directly shipped from the production barrier of Stanford VSC-approved vendors including Charles River Laboratories, Harlan, Taconic, and Jax.

Service scope and procedure:

Rederivation by Natural Mating Protocol

• Provide 5-7 males per line for heterozygotes or 3-5 males per line for homozygotes. Males should be 8-20 week old and proven breeders. Males should be housed individually at all times.

• Males should be mated with a test female, e.g., a young sexually mature ICR or CD-1 female, 5 – 7 days before the scheduled rederivation. This will establish the males as studs and increase the number of zygotes for the rederivation.

• Males are ideally left in your colony because space is limited in the Transgenic Facility. However, if your colony is off-campus, e.g., Arastradero, Blood Center, other arrangements will be made with the VSC.

• As a general rule, if your colony is located off-campus, allow two weeks for preparation of the mice before the actual date of the experiment. Two weeks prior to the experiment, while the males are still in your colony, set them up with females as described, to maximize the embryo yield. One week prior to the experiment, the mice will be transferred to RAF1 in order to allow for their acclimation and hormone treatment. For the transfer paperwork provide the location, room number, rack and shelf numbers, and the bar codes for the cages to be transferred. Once the mice are transferred to RAF1 they cannot go back to their original off-campus location.

• Provide 10 - 14 females per line at 4 - 5 weeks of age. If you are able to use commercially available females, this will greatly facilitate the rederivation. Females will be ordered, superovulated and mated with your stud males by Transgenic Facility personnel.

• It is very important to continue breeding your mice to obtain an additional 10 - 14 females in the event that the first rederivation attempt fails, or yield a low number of pups. This will allow the Transgenic Facility to quickly repeat the rederivation. Having sufficient females of the correct age is the limiting step in embryo transfer rederivation. The male pups in the litter may be culled at 3-4 days of age.

• It is strongly recommended that you genotype the pups, even if they are expected to be homozygotes, to ensure you have rederived your strain. If genotyping cannot be performed quickly in your lab, the use of a commercial genotyping service, e.g., Transnetyx, is recommended as it will be cost-effective in the long run.

Procedures for users:

1). Fill out and submit an online requisition form at http://med.stanford.edu/transgenic/form_embryoderivation.html. In the “Special Requirements” box, please list for each line the strain name (correct nomenclature, please), genotype, how many mice (indicating male or female) you will provide, and their ages. This information is needed to schedule the rederivation.

2). Confirm your rederivation date with Hong (hongzeng@stanford.edu)

3). Prepare males and female donors or tell us what strain of females to order if using wild-type females

4). If/when your mice are located on campus, contact Hong Zeng (hongzeng@stanford.edu), and arrange a meeting with one of the Transgenic Facility technicians. At the meeting, you will label the cages with the cards indicating "For Transgenic Facility". You should provide the Transgenic Facility with access information and key/card into your mouse room, as well as a sheet of bar code labels with your account number, to be used on the cages after the rederivation.

5). If your mice are located off-campus, they will be transferred to RAF1 one week prior to the experiment. Please provide the location, room number, rack and shelf numbers, and the bar codes for the cages to be transferred by emailing Hong (hongzeng@stanford.edu). If your test females are still with the males at the time of transfer, please inform the Transgenic Facility so that the mice can be separated.

 

Other protocols

Mouse colony expansion protocol

Rederivation by in vitro fertilization (IVF) protocol (service available soon)

Mouse reproductive biology for strain rederivation

 


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