The rat is one of the most commonly used experimental animal species in biomedical research. The availability of new research tools in rats could therefore provide considerable advances in the areas where this mammal is extensively used. We report the development of a new green fluorescent protein (GFP) rat strain suitable for organ transplantation and the birth of GFP rats following orthotopic transplantation of neonatal ovaries from this newly developed GFP rat strain to a wild-type Fischer 344 (F344) strain. A new GFP rat strain was developed by backcrossing eGFP Sprague-Dawley (SD-Tg(CAG-EGFP)Cz-004Osb) to wild-type F344 for eight generations. Whole ovaries from postnatal day (PND) 8 or PND 21 GFP rats were transplanted orthotopically to bilaterally ovariectomized wild-type adult females (n=6). All recipients were mated, and three of the five resulting litters contained GFP pups. In the PND 8 group, all recipients cycled regularly and the ovarian morphology appeared normal when collected at 9 months post-transplantation. In the PND 21 group, 60% of the recipients displayed regular estrous cycles at 9 months post-transplantation, but showed reduced ovarian size. This new strain and neonatal orthotopic transplantation could be useful for many biomedical fields including transplantation, development, and reproductive toxicology.