| The Yellow River carp(Cyprinus carpio haematopterus)has high protein content,rich nutrition,high edible value and commercial value,so it is an important freshwater fish culture species in China.Body color is one of the important economic traits of Yellow River carp.However,in recent years,abnormal body color has frequently occurred in the breeding process of Yellow River carp,which seriously affect the economic value of Yellow River carp.Therefore,it is urgent to explore the molecular genetic mechanism of Yellow River carp body color.In order to explore the molecular mechanism of abnormal body color of Yellow River carp,this study carried out genome-wide association analysis using normal(black)and abnormal(red)Yellow River carp as experimental materials.Based on the results of genome-wide association analysis,functional annotation and literature survey were conducted to screen out the key target gene s that affect the Yellow River carp body color.Then,the expression patterns of target genes were detected by real-time quantitative PCR technology.The full-length sequence of the target gene was obtained by RACE cloning,and its physicochemical properties,structural domains,and antigenic epitopes were predicted.The above research results can provide valuable data support for the follow-up related experiments.The main results of the study are as follows:(1)The pigment cells of normal and abnormal Yellow River carp in different periods were observed.The results showed that the body surface of normal Yellow River carp at larval stage was covered with radial melanocytes,but no melanocytes were observed on the body surface of abnormal Yellow River carp.The mature normal Yellow River carp contains yellow cells and two types of m elanocytes,while the abnormal Yellow River carp only has erythrophore.(2)Genome-wide association analysis was performed on normal and abnormal Yellow River carp,and 24,857 SNP loci related to pigment were obtained,including 18 SNP loci that were significantly related to body color.All genes in candidate genomic regions containing significant SNP loci were searched and a total of 202 differential genes were obtained.Five can didate genes,mitf,oca2,ap1m1,apoeb and lrp8,were obtained by functional annotation and literature research as possible candidates for involvement in the pigmentation process.Mitf,oca2 and ap1m1 are key genes for melanin synthesis,while apoeb and lrp8 may be involved in carotenoid metabolism.(3)The expression patterns of oca2-1 and oca2-2 genes were detected by real-time quantitative PCR.oca2-1 and oca2-2 were expressed in early stages,and the expression level of oca2-1 decreased significantly on the 7th day after hatching of abnormal Yellow River carp(p<0.05),while oca2-2 expression was not significant difference.Oca2-1 and oca2-2 showed tissue-specific expression patterns in adult fish.The expression of oca2-1 gene in the skin of abnormal Yellow River carp was significantly decreased(p<0.05)and was about 0.08 times that of normal Yellow River carp skin.There was no significant difference in the expression of oca2-2 in abnormal Yellow River carp skin.It is inferred that the oca2-1 retains more functions of the oca2 during the evolution of the Yellow River carp,and plays an important role in the formation of the body color of the Yellow River carp.(4)The full length of oca2-1 was obtained by RACE cloneing and sequence analysis was performed.The results showed that the full length of oca2-1 sequence was 2935 bp and the full length of CDS region was 2169 bp,encoding 722 amino acids.The oca2-1 is predicted to encode a hydrophobic protein and contains 13 transmembrane helix structures.The α helix is one of the most important components of the Yellow River carp OCA2-1 protein.Protein sequence homology analysis showed that the sequence homology between OCA2-1 and zebrafish OCA2 protein was as high as 95.8%. |
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