|Embryonic Stem Cell Lab||Transgenics & Knockouts||Cryopreservation Services& IVF|
|Histology & Pathology||Immunology Lab||Microscopy & Digital Imaging|
Contact: Dr. Rebecca Haffner
In a typical session, microinjected or aggregated embryos are transferred to 20 pseudopregnant females (at least 200 microinjected embryos). This should result in 50-70 live born offspring, with about 3 transgenic founders or chimeras. For transgenic mice, if there are no other specific requirements, the DNA will be injected into CB6/F1 zygotes and they will be transplanted into albino outbred females. For knockout mice, aggregations will be made between ES cells and outbred ICR embryos. Blastocyst injection will be performed if requested. The mice are kept at the unit till the offspring from the surrogate females are born. Thereafter, the TG or KO mice are transferred to the regular animal rooms and the department's maintenance and purchase costs apply.
Investigators wishing to generate a transgenic mouse are required do the following:
- When your construct is nearing completion submit an application for IACUC approval. Use the template for production of transgenic mice.
- Purify the construct exactly according to instructions on the site. Bring the purified construct to the unit, and place an order in Internal Services. If there are days which are not convenient for injection, this must be stated in the Notes section on the order form. Once the order is received you will be informed of the approximate date of birth of the pups, or the date for analysis for transient transgenics.
- BAC constructs – When your BAC construct is completed and is ready to be purified by PFGE, contact Golda Damari (ex3221) and place an order on Internal Services. You will be given a date to bring the DNA to the unit for injection. BAC DNA should be purified for injection using the NucleoBond® BAC 100 purification kit - see instructions.
All transgenic and gene targeting experiments must have prior IACUC approval.
DNA for pronuclear microinjection must be prepared according to the instructions below. DNA of poor quality or low concentration will not be injected.
For targeting experiments, ES cells prepared at the ES cell lab at the GMAF are preferred.