| The spotted scat(Scatophagus argus)is a popular cultured species in the southeast coast of our country.There are obvious differences in the growth of male and female spotted scat.The growth rate of females is faster than males.At the same time,spotted scat is used as ornamental fish due to the beautiful coloration in males.However,there are no reports on the breeding of unisexual seedlings.The genetic basis research is relatively weak,and the genome sequence is still missing,hindering the application of genome-assisted breeding of spotted scat.In this study,the genomic characteristics of spotted scat were evaluated by the second-generation sequencing technology,followed by the chromosome level reference genome with 99.73%coverage assembled by Pac Bio Hi-C sequencing.Based on the genome annotations,comparative genomics analysis was carried out to analyze some of the biological characteristics of the spotted scat adaptation to the environment.In addition,using the genomic data of male and female,one molecular marker was successfully developed that can quickly identify the genetic sex of spotted scat,which provides important technical support for the sex control breeding of spotted scat,and also provides a basis for the study of the sex determination mechanism of spotted scat.The main research results are as follows.1.Using Illumina sequencing technology to sequence the genome of one female and one male.According to K-mer analysis,it is estimated that the genomic size of spotted scat is about 600 Mb,the heterozygosity is 0.37%-0.38%,and the content of repetitive sequences is 26.99%-27.06%.In addition,the number and distribution characteristics of microsatellites in the genome were analyzed.Di-nucleotides duplication is the most abundant microsatellite type.The open reading frame sequence(ORF)of dmrt1 was compared to the male and female genomes.The result revealed that dmrt1 was only present in the male genomes,confirming that dmrt1 is a male-specific candidate gene for sex determination in spotted scat.2.The genome of female spotted scat was sequenced by Pac Bio and Hi-C sequencing techniques,and a 572.42 Mb reference genome was obtained with contig N50 21.05 Mb,scaffold N50 24.67 Mb,and 99.73%(570.88 Mb)of the sequences were attached to 24 chromosomes.BUSCO assessment of genomic integrity is about96.97%.A total of 24,256 protein-coding genes were predicted,among which 96.30%(23,359)of the genes were functionally annotated in the protein databases.The immune related gene family has been expanded by comparative genomic analysis of single copy homologous genes,such as immunoglobulin and major histocompatibility complex(MHC)families,which may be the genetic molecular basis of strong disease resistance of spotted scat.3.Based on the genomic data of male and female spotted scat,the gene sequences of Y chromosome specific dmrt1 and X chromosome specific dmrt1 b were compared and analyzed.According to the difference of sex chromosome sequences,one molecular marker that could quickly identify the genetic sex of spotted scat was successfully screened,and a method for rapid identification of the genetic sex of spotted scat was established.In summary,this study obtained a high-quality reference genome of spotted scat,confirmed that its sex-determining gene is the male-specific dmrt1 from the genome level,and developed a method for rapid identification of genetic sex of spotted scat,which not only provided an important technical support for molecular marker-assisted breeding,but also laid a foundation for the study of sex-determining genes and sex chromosome evolution of spotted scat. |
