Devising assisted reproductive technologies for wild-derived strains of mice: 37 strains from five subspecies of Mus musculus

PLoS One. 2014 Dec 3;9(12):e114305. doi: 10.1371/journal.pone.0114305. eCollection 2014.


Wild-derived mice have long offered invaluable experimental models for mouse genetics because of their high evolutionary divergence from laboratory mice. A number of wild-derived strains are available from the RIKEN BioResource Center (BRC), but they have been maintained as living stocks because of the unavailability of assisted reproductive technology (ART). In this study, we sought to devise ART for 37 wild-derived strains from five subspecies of Mus musculus maintained at the BRC. Superovulation of females was effective (more than 15 oocytes per female) for 34 out of 37 strains by treatment with either equine chorionic gonadotropin or anti-inhibin serum, depending on their genetic background (subspecies). The collected oocytes could be fertilized in vitro at mean rates of 79.0% and 54.6% by the optimized protocol using fresh or frozen-thawed spermatozoa, respectively. They were cryopreserved at the 2-cell stage by vitrification with an ethylene glycol-based solution. In total, 94.6% of cryopreserved embryos survived the vitrification procedure and restored their normal morphology after warming. A conventional embryo transfer protocol could be applied to 25 out of the 35 strains tested. In the remaining 10 strains, live offspring could be obtained by a modified embryo transfer protocol using cyclosporin A treatment and co-transfer of ICR (laboratory mouse strain) embryos. Thus, ART for 37 wild-derived strains was devised successfully and is now routinely used for their preservation and transportation. The information provided here might facilitate broader use and wider distribution of wild-derived mice for biomedical research.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animal Husbandry
  • Animals
  • Breeding / methods*
  • Cryopreservation*
  • Embryo Transfer
  • Female
  • Male
  • Mice, 129 Strain
  • Mice, Inbred BALB C
  • Mice, Inbred C3H
  • Mice, Inbred C57BL
  • Oocytes*
  • Reproductive Techniques, Assisted
  • Spermatozoa*

Grants and funding

The work was supported by a Grant-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan (No. 25112009 and No. 23220011 to AO) and by the RIKEN Epigenetics Program (Strategic Programs for R&D to AO). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.