Cryopreservation and transplantation of ovarian tissue in Japanese quail (Coturnix japonica)

Cryopreservation of avian genetic material is limited in terms of practicality, efficiency, and success. The complex structure of female gametes in birds makes the application of cryopreservation extremely difficult, and the fertilization obtained from frozen/thawed poultry semen is low and unpredictable. Embryonic germ cells can be frozen and used to generate germline chimeras, but the low efficiency and complex procedures limit these techniques for genetic conservation. The techniques of transplantation of ovaries and testes in newly hatched chicks have recently been developed and donor-derived offspring could be efficiently produced from cryopreserved testicular transplants and from fresh ovarian tissue transplants in chicken. This success in chicken gonadal transplantation provides an effective method for recuperating live birds from cryopreserved gonadal tissue. Research efforts are needed to develop cryopreservation technique for ovarian tissues and further apply cryopreservation and transplantation to the other avian species. The Japanese quail is a much smaller bird than the chicken and has a slightly different reproductive biology. Preliminary research has demonstrated that live offspring could be obtained from fresh ovarian grafts in one-week old quail. Using the Japanese quail as a model, the objective of this thesis is to develop a feasible and reliable cryopreservation technique for ovarian tissue and to recover it by subsequent transplantation, with the hope that this protocol of cryopreservation and transplantation can attribute to female germplasm conservation for other avian species in the future. Both slow-freezing and vitrification were used as cryopreservation approaches in this study. The efficiency was evaluated at three levels: cell viability, tissue histology and the recovery of the donor fertilities after surgical transplantation. Donor-derived offspring were obtained from the ovarian tissues that had been cryopreserved by either slow-freezing or vitrification. The vitrification protocol used in this study showed better outcomes at each level of evaluation. Thus the current study verified that the function of ovarian tissue in an avian species can be successfully preserved at subzero temperatures and recovered by transplantation. The vitrification protocol is recommended for future use concerning its low cost, high efficiency and overall simplicity in optimization to benefit more avian species.Land and Food Systems, Faculty ofGraduat