New Services

CRISPR/Cas9 based targeted transgenesis at H11 locus

The TMF now offers CRISPR / Cas9 mediated targeted transgenesis as a service.  The Hipp11 (H11) locus is a "safe-harbor" locus close to the centromere of mouse chromosome 11. Transgenes integrated into this locus are often expressed at a more consistent level than at the similar ROSA26 safe-harbor locus on chromosome 6.  An additional advantage is that a high efficiency of transgenesis (~ 50%) can be achieved at the H11 locus using CRISPR / Cas9 in conjunction with a plasmid based targeting construct of up to 10kb in size. Constructs can be easily inserted into the targeting vector using Gibson cloning. Please contact Jon Neumann for additional information. 

Improved Services

Hyper-ovulation of egg donors

To continue to address the 3R's and to reduce the cost of several of our services, the TMF is now using hyper-ovulation of female mice to produce oocytes and fertilized oocytes for different services. The difference between conventional superovulation and hyperovulation is the inclusion of an antibody against inhibin, which is administered along with the PMSG. During superovulation, exposure to FSH induces synthesis of inhibin which inhibits secretion of endogenous FSH from the anterior pituitary. Addition of antisera against inhibin blocks this process, resulting in increased numbers of oocytes being ovulated per mouse. For example, during re-derivation via IVF, the TMF can reduce the number of females used as egg donors from 12-14 down to 5 - 6, a 50% reduction. This reduces the number of animals requried for this, and related, processes and also helps to reduce costs of services. We obtain our hyperovulation reagents form Cosmo Bio Ltd. See Takeo & Nakagata (2015) PLoS One 10: e0128330 for more details. 

Innovative Strategies

Production of sex-sorted blastocysts

By mating wildtype females with male mice carrying an X-linked GFP transgene, we can produce blastocysts that can be sorted by sex (female embryos are fluorescent but male embryos are not).  When such blastocysts are injected with ES cells to make chimeric mice and the resulting pups are sexed at birth, we can immediately see if any sex conversion has taken place (e.g., by male ES cells being injected into female embryos, or vice versa), which is a strong indication of germline capability.

identification of female embryos using X-linked GFP transgene

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