Successful Production of an All-Female Common Carp (Cyprinus carpio L.) Population Using cyp17a1-Deficient Neomale Carp

Due to sexual dimorphism in the growth of certain cultured fish species,the production of monosex fishes is desirable for the aquaculture industry.Nowadays,the most widely practiced technique available for the mass production of monosex fish populations is sex steroid-induced sex reversal.Here,a novel strategy for the successful production of all-female(AF)common carp(Cyprinus carpio L.),to take advantage of the sexual dimorphism in growth documented in this species,has been developed using genetic engineering via single gene-targeting manipulation without any exogenous hormone treatments.Male and female heterozygous cyp17a1-deficient common carp were first obtained using the clustered regularly interspaced short palindromic repeats/CRISPR-associated endonuclease 9(CRISPR/Cas9)technique.An all-male phenotype for homozygous cyp17a1-deficient carp,regardless of the individuals’sexdetermination genotypes(XY or XX),has been observed.A male-specific DNA marker newly identified in our laboratory was used to screen the neomale carp population with the XX genotype from the cyp17a1-deficient carp.These neomale carp develop a normal testis structure with normal spermatogenesis and sperm capacity.The neomale common carp were then mated with wild-type(WT)females(cyp17a1^(+/+)XX genotype)using artificial fertilization.All the AF offspring sample fish from the neomale-WT female mating were confirmed as having the cyp17a^(1+/-)XX genotype,and normal development of gonads to ovaries was observed in 100.00%of this group at eight months post-fertilization(mpf).A total of 1000 carp fingerlings,500 from the WT male and female and 500 from the neomale and WT female mating,were mixed and reared in the same pond.The average body weight of cyp17a1^(+/-)XX females was higher by 6.60%(8 mpf)and 32.66%(12 mpf)than that of the control common carp.Our study demonstrates the first successful production of a monosex teleost population with the advantages of sexual dimorphism in growth using genetic manipulation targeting a single locus.