The integrity of a GEM facility is dependent upon the ability to maintain the mouse colony in a specific pathogen free (SPF) state. Therefore, when Baylor College of Medicine Investigators desire to import mouse strains from facilities that are not SPF or when an outbreak of contamination occurs in a mouse colony, the GERM Core will help investigators to rederive their mouse strains using embryo transfer to recipient SPF female mice.
What Will Happen
- Investigator will submit a request form through iLabs, filling out detail information on male and female mouse numbers and ages. An account number and IACUC protocol number must be provided at this time. The Investigator IACUC protocol must be approved for all procedures performed in the user laboratory.
- The core will review investigator provided information and approve the request. The core will schedule the colony expansion according to the female mouse age and coordinate with the investigator or CCM if the mice are in quarantine.
- Investigators will complete steps 3-10. Investigator prepares 2+ adult males and 2-4+ young female mice at 3-5 weeks of age and of the appropriate genotypes.
- One week before the scheduled rederivation date, pick up 1 tube of M2 media, 1 tube of PMSG, and 1 tube of HCG from the GERM Core. Note: Bring an ice bucket to put the hormones in and put them in a -20C freezer when you get back to your lab. Dilute the hormones right before you go to give the injections.
- PMSG is given at 10 a.m. three days before the scheduled rederivation date (e.g. Tuesday for cryopreservation on Friday). Dilute the PMSG with 950 ul of 0.9% saline and administer 0.1cc, i.p. per mouse.
- HCG is given 47 hours after PMS at 9 a.m., one day before the scheduled rederivation date (e.g. Thursday for procedure on Friday). Dilute the HCG with 950 ul of 0.9% saline and administer 0.1cc, IP per mouse.
- Place the females with the males immediately after administering HCG.
- The morning following the HCG injections, the females are removed from the male cages and oviducts are collected into M2 media provided by the GERM Core (a protocol will be provided).
- Label the tubes with the request number generated by iLab. Deliver the oviducts to R119A and leave in the biosafety hood no later than 9 a.m.
- The core will dissect, digest, and collect all oocytes from the oviducts, identify and transfer all fertilized 1-cell embryos to pseudo pregnant recipient ICR female mice to allow gestation.
- Live-born founder animals will be held by the GERM Core until 10-14 days of age and subsequently transferred to the User Investigator.
What is Expected
- The GERM core will transfer all 1-cell embryos to 0.5-day pseudo pregnant recipient mice.
- The number of expected live-born offspring will vary and will be dependent on the number of embryos transferred. On average, 15-20% of transferred embryos result in live-born offspring.
- If there are no fertilized embryos collected, the core will charge 40% of cost. Investigators can repeat the cryopreservation at 100% cost.