Mating Schemes to generate 2-cell embryos for cryopreservation.
The approximate goal for each line is to freeze at least 200 embryos @20 embryos/vial. To obtain this many embryos in a single day females must be induced to superovulate prior to mating. Due to variability in response to superovulation regimens and breeding performance, however, it may take more than one attempt to collect enough embryos. Depending on the mating pairs chosen, not all the embryos will necessarily carry the gene (transgene or knockout allele) of interest. For example, in the case of heterozygous mutant males mated with wildtype females, only 50% of the resulting pups will carry the gene of interest. This means that after freezing and thawing, the pups derived from frozen embryos will have to be genotyped and additional breeding performed to reconstitute the line.
The following is a guideline only; it is important to discuss specific details for your paricular line with the TPS Director well in advance.
You must provide us with 10-12 heterozygous or homozygous transgenic (Tg) or knockout (KO) males at 8 weeks to 8 months of age. When selecting males, make sure they have not been used for breeding for at least 2 weeks and have been housed individually (1/cage) for about two weeks. We will also need 10-12 embryo-donor females of superovulatory age (10-12 grams). Wildtype females will be purchased for this purpose.
Superovulation will be induced by injecting the gonadotropins, PMS (pregnant mare's serum), followed by HCG (human chorionic gonadotropin) 48 hrs later. Superovulated mice will be mated with the males immediately after the second injection. Two days later, two-cell embryos will be collected from plugged females and frozen.
If your animals are housed in the BRL (Biologic Resources Laboratory), hormone injections, mating, and dissection of oviducts from plugged females will be performed by BRL staff. The oviducts will be transported to our facility for embryo collection/freezing.
Please note, all animal purchases /per diem will be charged to your BRL account.
