Silver arowana, Osteoglossum bicirrhosum, is sexually monomorphic, and its sex determination mechanism is poorly understood, which has been a major hurdle in its captive breeding. To this end, we assembled two high-quality chromosome-level genomes for each sex of silver arowana, with chromosome N50 of 10.6 Mb and 10.4 Mb in female and male individuals, respectively. Combing re-sequencing data of 109 individuals, we identified two sex-specific SNPs and confirmed a ZW model of sex-determination in this species. Both sex-specific SNPs are located in about 26Kb upstream of foxl2 gene. Since foxl2 has been confirmed to be differentially expressed in the gonads of silver arowana, it probably plays an important role in sex determination via the cross-regulation with auxiliary genes significantly correlated to it, such as dmrt1, sox8a and cyp11a1, etc. The sex chromosomes are homomorphic with a potentially recent origin, as linkage disequilibrium analysis showed minor recombination suppression. Overall, we developed accurate molecular markers for sex identification of silver arowana in each developmental stage, subsequently improving the reproductive efficiency and further promoting the industrial development.