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Genetic introgression of escaped farmed Atlantic salmon ( Salmo salar ) into wild populations is a major environmental concern for the salmon aquaculture industry. Using sterile fish in commercial aquaculture operations is, therefore, a sustainable strategy for bio-containment. So far, the only commercially used methodology for producing sterile fish is triploidization. However, triploid fish are less robust. A novel approach in which to achieve sterility is to produce germ cell-free salmon, which can be accomplished by knocking out the  dead-end  ( dnd ) gene using CRISPR-Cas9. The lack of germ cells in the resulting  dnd  crispants, thus, prevents reproduction and inhibits subsequent large-scale production of sterile fish. Here, we report a rescue approach for producing germ cells in Atlantic salmon  dnd  crispants. To achieve this, we co-injected the wild-type (wt) variant of salmon  dnd  mRNA together with CRISPR-Cas9 constructs targeting  dnd  into 1-cell stage embryos. We found that rescued one-year-old fish contained germ cells, type A spermatogonia in males and previtellogenic primary oocytes in females. The method presented here opens a possibility for large-scale production of germ-cell free Atlantic salmon offspring through the genetically sterile broodstock which can pass the sterility trait on the next generation.

Rescue of germ cells in dnd crispant embryos opens the possibility to produce inherited sterility in Atlantic salmon