Onychostoma macrolepis is an emerging commercial cyprinid fish species. It is a model system for studies of sexual dimorphism and genome evolution. Here, we report the chromosome‐level assembly of the O.macrolepis genome obtained from the integration of nanopore long‐read sequencing with physical maps produced using Bionano and Hi‐C technology. A total of 87.9 Gb of nanopore sequence provided approximately 100‐fold coverage of the genome. The preliminary genome assembly was 883.2 Mb in size with a contig N50 size of 11.2 Mb. The 969 corrected contigs obtained from Bionano optical mapping were assembled into 853 scaffolds and produced an assembly of 886.5 Mb with a scaffold N50 of 16.5 Mb. Finally, using the Hi‐C data, 881.3 Mb (99.4% of genome) in 526 scaffolds were anchored and oriented in 25 chromosomes ranging in size from 25.27 to 56.49 Mb. In total, 24,770 protein‐coding genes were predicted in the genome, and ~96.85% of the genes were functionally annotated. The annotated assembly contains 93.3% complete genes from the BUSCO reference set. In addition, we identified 409 Mb (46.23% of the genome) of repetitive sequence, and 11,213 non‐coding RNAs, in the genome. Evolutionary analysis revealed that O. macrolepis diverged from common carp approximately 24.25 million years ago. The chromosomes of O. macrolepis showed an unambiguous correspondence to the chromosomes of zebrafish. The high‐quality genome assembled in this work provides a valuable genomic resource for further biological and evolutionary studies of O. macrolepis.

Dnmt3a, a de novo methyltransferase, is essential for mammalian germ line DNA methylation. Only one Dnmt3a is identified in mammals, and homozygous mutants of Dnmt3a are lethal, while two Dnmt3a paralogs, dnmt3aa and dnmt3ab, are identified in teleosts due to the third round of genome duplication, and homozygous mutants of dnmt3aa and dnmt3ab are viable in zebrafish. The expression patterns and roles of dnmt3aa and dnmt3ab in gonadal development remain poorly understood in teleosts. In this study, we elucidated the precise expression patterns of dnmt3aa and dnmt3ab in tilapia gonads. Dnmt3aa was highly expressed in oogonia, phase I and II oocytes and granulosa cells in ovaries and spermatogonia and spermatocytes in testes, while dnmt3ab was mainly expressed in ovarian granulosa cells and testicular spermatocytes. The mutation of dnmt3aa and dnmt3ab was achieved by CRISPR/Cas9 in tilapia. Lower gonadosomatic index (GSI), increased apoptosis of oocytes and spermatocytes and significantly reduced sperm quality were observed in dnmt3aa−/− mutants, while normal gonadal development was observed in dnmt3ab−/− mutants. Consistently, the expression of apoptotic genes was significantly increased in dnmt3aa−/− mutants. In addition, the 5-methylcytosine (5-mC) level in dnmt3aa−/− gonads was decreased significantly, compared with that of dnmt3ab−/− and wild type (WT) gonads. Taken together, our results suggest that dnmt3aa, not dnmt3ab, plays important roles in maintaining gametogenesis in teleosts.

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Background: Dnmt3a , a de novo methylatransferase, is essential for both male and female germ line DNA methylation. Only one Dnmt3a is identified in mammals, and homozygous mutation of Dnmt3a is lethal, while two Dnmt3a , dnmt3aa and dnmt3ab , are identified in teleosts due to the third round of genome duplication, and homozygous mutation of dnmt3aa and dnmt3ab is viable in zebrafish. Dnmt3aa and dnmt3ab were demonstrated to have essential and non-overlapped functions on modulating behavioral control, however, their function in gonadal development is unclear in fish. Results: In this study, the expression patterns of dnmt3aa and dnmt3ab in developing gonads of Nile tilapia was analyzed by quantitative real time PCR and fluorescence in situ hybridization. Both dnmt3aa and dnmt3ab displayed sexually dimorphic expression in developing gonads. Dnmt3aa was widely expressed in gonadal germ cells and somatic cells, highly expressed in oogonia, phase I and II oocytes and granulosa cells in ovaries and spermatogonia and spermatocytes in testes, while dnmt3ab was mainly expressed in ovarian granulosa cells and testicular spermatocytes. Mutation of dnmt3aa and dnmt3ab was achieved by CRISPR/Cas9 in tilapia. Lower GSI (Gonadosomatic index), increased apoptosis of oocytes and spermatocytes and significantly reduced sperm quality were observed in dnmt3aa −/− mutants, while no obvious phenotype was observed in dnmt3ab −/− mutants. Consistently, the expression of apoptotic genes was significantly increased in dnmt3aa −/− mutants. In addition, dnmt3aa and dnmt3ab were found to have certain compensatory effects in the gonads. The global DNA methylation level in ovaries and testes of dnmt3aa −/− mutants was decreased significantly, compared with that of dnmt3ab −/− mutants and WT. Conclusions: Taken together, our results suggest that dnmt3aa , not dnmt3ab , plays important roles in maintaining gametogenesis in teleost. Our results enrich the understanding of the function of DNA methyltransferases in gonads of non-mammalian vertebrates.