Comparative Analysis Of DNA Methylation In Muscle Tissues Of Takifugu Rubripes,Takifugu Obscurus And Reciprocal Cross Puffer

Puffers is one of the most economically valuable fish species in East Asia and has been extensively cultured in China,Japan,and Korea.Among them,Takifugu obscurus and Takifugu rubiripes are the main breeds cultured,which are also the only two species allowed for market sale in China.Takifugu rubripes has a larger body size and grows faster than other genus of Takifugu.Compared to Takifugu rubripes,Takifugu obscurus is smaller in body size,grows slower,but exhibits better meat quality and mouthfeel,making it one of the famous " three gastronomies in the Yangtze River ".Artificial propagation and aquaculture management of puffers has been established over the past few decades.However,little progress has been made in the cultivation and improvement of puffers species.To breed a new species that can combine the growth advantage of Takifugu rubripes and the good meat mouthfeel of Takifugu obscurus,we have artificially bred the reciprocal cross puffer with Takifugu rubripes as the mother and Takifugu obscurus as the father.To explore the differences in growth among the three species,the Takifugu rubripes,Takifugu obscurus,and reciprocal cross puffer,we analyzed muscle tissue for differences in DNA methylation levels and distinctions in methylation patterns using whole genome bisulfite sequencing(WGBS).Clean reads of Takifugu rubripes23240690,Takifugu obscurus 19484211,and reciprocal cross puffer 19928156 were obtained,totaling 626653057 clean reads,after bioinformatic means analysis.The methylation pattern was dominated by the CG type,and the percentage of CG sequences was around 61%,which was higher than that of the CHG and CHH types.The go enrichment data processing analysis identified significant enrichment in the following categories: regulation of biological process,cell signaling,cell communication,transcription regulator activity,DNA binding transcription factor activity,membrane.These biological processes,molecular functions,cell composition were enriched in DMR associated genes.Thus indicating that DNA methylation modifications are widely present in these life activities.Data processing analysis of KEGG enrichment data revealed that DMR associated genes were significantly enriched in the following pathways: neuroactive ligand receptor interaction,MAPK signaling pathway vascular smooth muscle contact,calcium signaling pathway,endocytosis.Contrasting the differential genes in the three screened cohorts revealed relatively distinct DNA methylation modification differences in bone morphogenetic proteins(BMPs),myogenic regulatory factors(MRFs),paired box transcription factors(pax).It is speculated that the hypermethylation of gdf6 a gene inhibits the formation and development of cartilage and bone of Takifugu obscurus.Hypermethylated modification of Myf5 gene,inhibits proliferative differentiation of myoblasts at the initial stage of Takifugu obscurus embryonic development.Hypermethylated modification of the Myo G gene that inhibits the conversion of Takifugu obscurus myocytes to muscle fibers.Hypermethylation of pax3 b,and pax7 a,inhibits the growth and development of muscle satellite cells in Takifugu obscurus.Results in a significantly lower growth advantage in the Takifugu obscurus compared to Takifugu rubripes and reciprocal cross puffer when multiple genes are combined.The reciprocal cross puffer,because of the combined growth advantage of the Takifugu rubripes,whose growth velocity is significantly elevated compared with that of the Takifugu obscurus.The extent of methyla-tion modification was also markedly decreased in the above-mentioned genes,leading to a clear increase in their gene expression,as reflected by heterosis in reciprocal cross puffer.This study analyzed DNA methylation differences in the Takifugu rubripes,Takifugu obscurus and reciprocal cross puffer,and explored whether these differences are associated with heterosis in the reciprocal cross puffer.It provides a theoretical basis and reference basis for the future cultivation and improvement of new cultivars of reciprocal cross puffer.