Study On The Genetic Mechanism Of The Skin Color Mutation In Yellow River Carp And Russian Sturgeon

Skin color is crucial for fish farming and is an important indicator for its commercial value.For ornamental fish,bright colors can increase the selling price,while for cultured fish,the appearance of abnormal commercial fish will reduce the value.Earlier researches on fish skin coloration were mainly based on traditional genetic methods such as observational phenotypes and crossbreeding.Since the emergence of high-throughput sequencing technologies,studies on fish skin color have been got preliminary results from the view of development and molecular genetics.The study of fish skin color in China mostly focuses on some color-rich varieties,such as the Koi carp and the Oujiang color carp.The research on the genetic mechanism of skin color of the Yellow River carp and Russian sturgeon is almost blank.In this study,we used genome-wide sequencing and transcriptome sequencing to study two different fish skin color samples,and analyzed the data on the basis of bioinformatics.Through processing data with Plink,Popoolation2,BWA,SAMtools,Trinity,Blast and other major software,we obtained pigment synthesis related genes.In the skin color study of Yellow River carp,based on the genomic data analysis method we used the carp high-throughput SNP genotyping chip for the red and black populations of the Yellow River carp to conduct the whole gene association analysis(GWAS).Then we used the Plink’s logistic regression model and Bonferronit’s statistical test to find the significant sites of the differences scanning SNPs.A total of 34,124 SNPs was obtained,and 189 functional genes were located in the reference genome,but there were no genes associated with pigment synthesis.The next step was to perform genome-wide sequencing of the two populations,and a total of 610,997,044 reads were obtained after cleaning up.Sequencing data were analyzed using BSA technology.The sequencing results of the two populations were first compared to the reference genome of carp,even the lowest alignment rate was 95%.The results of the alignment were synchronized using Popoolation2 and allelic frequency differences were calculated.After Fisher’s exact test,the most significant sites(-logP values greater than 8)15,016 were screened and 513 functional genes were mapped.Five genes related to melanin synthesis were found in the search,among these genes there were three genes: MITF was responsible for the transcription of tyrosinase;CREB1 was responsible for the transcription of MITF;ASIP inhibited eumelanin and promoted the formation of pheomelanin;the other two genes were expressed the cell membrane protein GNAS and OCA2,the former coupled with MC1 R activate CREB1 and MITF to turn on transcription of Tyr,the latter one assists the melanin synthesis substrate into melanosomes.This study suggests that the process of melanin synthesis may be inhibited in the red Yellow River carp compared to the black population and the pigment synthesis is dominated by pheomelanin in red population.In the second part we conducted a comparative transcriptome analysis in two skin kinds of Russian sturgeon(Acipenser gueldenstaedti),normal(black)phenotype and albino(withe)phenotype.Through sequencing on three pooling sample,total 277,268 of contigs were obtained as our reference sequences.The assembled sequences were aligned and annotated with the three major protein libraries,and yielded 32,965 genes.In the analysis of differential gene expression between these two groups,a total of 785 genes were found to be significantly different expressed.There were 400 genes up-regulated in the normal sample and 385 genes up-regulated in the albino sample.After that,16 genes were randomly selected for quantitative RT-PCR verification,and the results were consistent with the transcriptome expression levels.In the Gene functional category(GO),17,058 genes were annotations,including all differential genes.In the GO enrichment analysis,there were 35 pigment synthesis-related genes;Three of the 148 pathways for differential gene matching in the KEGG pathway analysis were associated with melanin synthesis.Combined with literature searches and data from this study,we have targeted nine genes including Dct,Tyrp1,SLC45A2,MC1 R,CNTS,PME1 a,PMElb,CD63 and BLOC1S3.Among the expression products of these genes,three are key enzymes in the process of catalyzing melanogenesis by using tyrosine as a substrate,two are for regulating the pH of melanosomes,three are for the formation of black bodies,and one is for transporting enzymes required by melanin synthesis.Combine literature search and our data,the study result proposes that the albino Russian sturgeon may have defects in generation of eumelanin and melanosomes contrasting to normal samples.Collectively,our findings provide genomic evidence for the evolutionary and genetic studies of the skin color variation of the Yellow River Carp and Russian sturgeon,and provide molecular genetic basis for the selective breeding of these two species of fish.