Analysis Of Female-biased Sexual Size Dimorphism And Sex-related Genes Cyp19a1 And Dmrt1 In Yellow Perch(Perca Flavescens)

Yellow Perch(Perca flavescens),a common cold water and fresh water fish widely distributed in North America.It is one of the most valuable 76 commercial fishing fish in the Great Lakes region of North America.Yellow Perch has the characteristics of,low fat,phospholipid and high protein content,strong disease resistance and wide adaptability,and it can tolerate high breeding density and has high breeding value.Because the female individuals of Yellow Perch have sex and size dimorphism,and the growth rate of female individuals is faster than the male fishes,and it is difficult to distinguish their gender outside the breeding period.Our research group cooperates with the Ohio State University to study and develop the unisexual cultivation of female Yellow Perch,and use transgenic perch to cultivate neo male population and rapidly growing unisexual female population to improve the growth performance of Yellow Perch.In order to understand how sex-related genes and mi RNAs regulate the sex determination mechanism of Yellow Perchs and develop markers for rapid identification of male and female,this paper screened sex differentially expressed and specifically expressed genes and mi RNAs through genome data and small RNA transcriptome analysis of Yellow Perch,used si RNA technology to RNA interfere with sex-related genes cyp19a1 and dmrt1,and screened sex-specific markers,SSR(simple repeat)and SNP(single nucleotide point mutation)for understanding the sex determination mechanism of Yellow Perch.The results are as follows:1.In order to further understand the differences in gene expression of female biased size dimorphism and the regulation mechanism of sex-related mi RNA of Yellow Perch,this study combined the whole genome of Yellow Perch which completed by our research group and the carried out the small RNA transcriptome sequencing for data processing.In this study,the differentially expressed genes and sex-specific genes of large female fish,small female fish and male fish were predicted,and enrichment analysis was carried out.By annotating micro RNAs,highly homologous mi RNAs and unknown specific mi RNA sequences were obtained,and the function of mi RNAs,the interacting target genes and the involved signal pathways were predicted.At the same time,we combined the genome and transcriptome to compare the specific sequences of sex chromosomes to screen the sequences specifically expressed,male specific marker and SNP.The analysis results of biased size dimorphism of Yellow Perch showed that there were significant differences in gene expression between large females and small females compared with males,and the trend of differential gene expression of small females was closer to males rather than large females.KEGG enrichment results showed that golden perch was mainly involved in three signal pathways and the number of genes involved were cancer pathway signal pathway(329),PI3K-Akt signal pathway(207)and MAPK signal pathway(202).In addition,CYP19(P450)involved in signal pathways were cytochrome P450 metabolic pathway of exogenous substances(11)and drug cytochrome P450 metabolic pathway(8).The mi RNA screening results showed that compared with the mi RBase database,it was found that there were 11 pairs of unique mi RNA sequences and 293 pairs of functionally conserved mi RNA sequences.2.In order to analyze the genetic resources and diversity of golden perch and screen out specific markers that can quickly identify the sex of Yellow Perch,RAD-seq was performed on 8 strains(state)of Yellow Perch in North Atlantic coastal(NY)、 South Atlantic coastal(NC)、 South Atlantic coastal(PA)、 Northwest Lake Plains(NE)、 Great Lakes watershed(OH)、Great Lakes watershed(WI)、Choptank and Perquimans.The results showed that the nucleotide diversity of Yellow Perch was 0.00303,and the results showed that 59766 SSR sequences were screened,including 49052 dinucleotide repeats(82.1%),6299 trinucleotide repeats(10.5%),3960 tetranucleotide repeats(6.6%),314 pentanucleotide repeats(0.5%)and 141 hexanucleotide repeats(0.2%).Based on the analysis of male specific marker and SNP by RAD-seq data,combined with the whole genome and transcriptome data,115 male specific marker and 9251 SNP were obtained,including 2457 single nucleotide mutation SNP and 1833 single nucleotide deletion mutation SNP.3.In order to better study the regulation mechanism of sex determining genes of Yellow Perch,this study selected female related gene cyp19a1 and male related gene dmrt1 for RNA interference to understand whether these two genes are the key genes of sex determination of Yellow Perch and whether these genes would affect the growth rate while affecting gender development..si RNA was synthesized by chemical modification.Four rounds of si RNA and si RNA simulant(negative control)were injected intraperitoneally to interfere with cyp19a1 and dmrt1 genes at the age of 51 to 121 days.The results showed that the female fish interfering with cyp19a1 gene could not reverse sex to male,and the gonads developed into intermediate sex,reduced the growth rate,gonadal development index GSI and increased liver weight index HSI.In the experiment of interfering with dmrt1 gene,it was found that some male Yellow Perch sexual reversal was female,which showed that the proportion of female fish increased from 45-55% to 65-70%.The GSI of female fish remained unchanged and the HSI increased,while the male fish without sexual reversal showed slow gonadal development,decreased GSI and increased HSI.Feeding low-dose MT feed(20 mg / kg)or high-dose MT feed(50 mg / kg)on 38 days can make 100% sexual reversal of females without intermediation.Therefore,it is concluded that the key time for the initiation of sex determination mechanism of Yellow Perch is 38-50 days old.