Alternative Approaches for Repressible Transgenic Sterilization of Channel Catfish (Ictalurus punctatus)

Abstract

Transgenic sterilization has the potential for accomplishing 100% reproductive confinement to avoid genetic communication of transgenic or domestic genotypes with wild populations. Two strategies of transgenic sterilization were developed and tested in channel catfish (Ictalurus punctatus). For the first strategy, three sterilization constructs (Nanos-nanos, Nanos-dnd, Dazl-vasa) were introduced into channel catfish embryos to overexpress the Bax gene specifically in the germ line to ablate the germ cells, leading to infertility. Four-year-old Nanos-nanos, Nanos-dnd, or Dazl-vasa construct exposed males, but not females, had significantly lower (p < 0.05) gonadosomatic index (GSI) than the control, and lower (p < 0.05) percentages (54.2%, 44.4%, 56.7%, respectively) of individuals with a gonad development score ≤ 2 (maximum = 5) than the control (0.0%). Mosaicism of transgene integration was widely observed in the P1 fish. Unexpectedly, doxycycline treatment did not effectively suppress expression of the transgene, indicating efficient reversible transgenic sterilization will likely not be possible for this system. This study demonstrated that the overexpression of Bax gene could lead to the death of germ cells in the male fish with high efficacy, and could be useful to produce sterile fish when repressible control is not needed. The second strategy aimed to disrupt the reproductive endocrine regulation of channel catfish by overexpression of glutamic acid decarboxylase (GAD) to increase γ-aminobutyric acid (GABA) level, thereby producing infertile fish. Repressed reproductive performance was observed in all generations of GAD transgenic fish examined, as revealed by less mature fish in the F1 generation at the age of 5, fewer fish spawned naturally in the F1 (6 and 9-year-old) and F2 (3-year-old) generations, and lower serum GnRH levels in the 1-year-old F2 fish. Interestingly, male transgenic fish showed lower (p < 0.05) serum levels of GnRH and testosterone than control fish at the age of 4, indicating a different regulation mode of GABA on GnRH in adult males than females as transgenic and control females were not different. Overexpression of GAD could repress the reproductive performance of channel catfish, hormone therapy could sometimes restore fertility, but further research is needed to make this approach 100% effective.