Genome-wide Association Analysis Of Sheep Meat Quality Traits And Functional Research Of Candidate Genes In Intramuscular Fat

With the improvement of people’s living standard,the requirement of quality mutton is higher and higher,so the breeding of sheep meat quality traits has been paid more and more attention.Meat color,marbling,pH,shear force,cooked meat rate,water loss rate,water content and intra muscle fat were closely related to the quality of mutton.SNPs significantly related to the above meat quality traits were obtained by genome-wide association analysis（GWAS）,and the candidate major genes were initially screened,and the functional studies of the candidate major genes were conducted.The aim is to provide scientific basis for improving the quality of mutton.Purpose:In this study,Texel sheep×Altay sheep F₂ was used to design the resource population,and462 individuals were genotyped using high-density SNP chips and whole genome resequencing,and whole-genome association analysis was conducted for sheep meat quality traits.Furthermore,the function and mechanism of JAZF1 gene annotated by SNP sites significantly related to intramuscular fat were studied.Methods:1.SPSS19.0 was used to analyze the phenotype data,normal distribution analysis and principal component analysis of 11 meat quality traits,including meat color,marbling,pH_45min,pH_24h,pH_48h,pH_72h,shear force,cooked meat rate,water loss rate,water content and intra muscle fat.2.Illumina sheep high density（600K）SNP chip and whole genome resequencing were used for genotyping of SNP loci distributed in the genome.Through the mixed linear model constructed by GEMMA software,genome-wide association analysis was conducted on the above 11 meat quality traits by using SNPs distributed on chromosomes after quality control,and SNPs significantly correlated with the target traits were screened.According to the sheep genome（Ovis＿aries＿v3.1）,the significant related SNPs found in GWAS were annotated,and the candidate genes were identified by gene function analysis.3.In order to explore the mechanism of JAZF1 gene on lipid differentiation,RNAi and overexpression of JAZF1 gene were used to detect lipid content by triglyceride（TG）.qRT-PCR and Western blot were used to detect the mRNA expression levels and protein expression levels of fat synthesis genes FAS,ACC,and fat decomposition genes ATGL and HSL,in order to analyze the influence of JAZF1 gene on lipid differentiation.Results:1.Phenotypic statistical analysis of 11 meat quality traits showed that the phenotypic data of each trait met the requirements of subsequent experimental data analysis.2.A total of 85 SNPs significantly correlated with the target traits were screened out by GWAS.The 85 SNPs were annotated according to the sheep genome（Ovis＿aries＿v3.1）,and 24 candidate genes were identified by gene function analysis,namely MEGF9 and GK2.The marbling candidate gene was TWISTNB gene;pH_45min candidate gene was CCDC92 gene;pH_24h candidate genes were KDM4C,GOT2 and TGFB2 genes.The pH_48hcandidate genes were MMP12 and MMP13.Candidate genes for pH_72h were HILPDA and FAT1.Shear force candidate genes were PDE1A,EREG and AREG genes.Candidate genes for cooked meat rate were DOCK4,AOAH and CREB5 genes.Candidate genes for water loss rate were PRKCE and PRKG1.Water candidate genes were ELOVL2,ADRA2A and RASGEF1A.The candidate genes of intramuscular fat were JAZF1 and SMAD1.3.By studying the function of JAZF1 gene,it was found that lipid content was significantly increased when JAZF1 gene was inhibited;FAS and ACC showed an increasing trend in transcription and protein levels,while ATGL and HSL showed a decreasing trend in transcription and protein levels.JAZF1 gene overexpression significantly decreased lipid content.FAS and ACC showed a decreasing trend in transcription and protein levels,while ATGL and HSL showed an increasing trend in transcription and protein levels.Conclusions:In this study,the F₂ resource population of 462 Texel sheep×Altay sheep was constructed,and 11 phenotypic databases of meat quality traits were established.Through statistical analysis of the phenotypic data,it was determined that intra muscle fat was the key phenotypic trait in the improvement of meat quality traits.By using 600K high density microarray and whole genome resequencing technology for genotyping,JAZF1 and SMAD1 genes were preliminarily identified as candidate genes affecting intramuscular fat by GWAS method,which provided an important reference for the exploration of functional genes for meat quality traits of sheep.JAZF1 gene RNAi and overexpression verification tests showed that JAZF1 gene played a significant role in the regulation of lipid differentiation,and JAZF1 gene could be an important candidate gene to improve lipid deposition in sheep.