Germ cell transplantation techniques have broad applications, including the production of live fish from cryopreserved germ cells, the conservation of endangered fish species, and the production of hybrid fish. However, no studies have explored interspecific germ cell transplantation between closely related species, such as White Crappie (WC) and Black crappie (BC).
This study investigates the efficiency and viability of ovarian germline cells (OGCs) transplantation from diploid WC into the triploid BC female to advance the production of hybrid crappie using a xenogenic method. The objective of this study was to assess the viability, integration, and proliferation of transplanted freshly prepared and one-month cryopreserved OGCs in the gonad of the triploid BC females. OGCs were isolated from age-1 diploid WC females, tagged with PKH-26 green-fluorescent dye, and transplanted into triploid BC females using a catheterization procedure. First, freshly prepared OGCs were first transplanted into one group of nine triploid BC females. Next, OGCs were cryopreserved for one month in liquid nitrogen using a Recovery™ cell culture freezing medium. The survivability rate of the cryopreserved OGCs was 80.28%. After one-month, cryopreserved OGCs were transplanted into a second group of nine triploid BC females. The presence and viability of the implanted OGCs were checked using a BD AccuriTM C6 Plus Flow Cytometer by randomly sacrificing three fish each at 15, 30, 45-days post-implantation.
Flow cytometry results indicated that both freshly prepared and cryopreserved transplanted OGCs were able to integrate and proliferate in the gonads of the host triploid BC females, creating a xenogenic fish. These findings suggest that OGCs transplantation between diploid WC and triploid BC female can be viable technique for producing xenogenic crappie for the production of hybrid crappie. The results also demonstrated the feasibility of long-term cryopreservation of OGCs for the hybrid crappie production and the conservation of female genetic material.