| With the increasing improvement of residents’ living standards,people’s dietary structure and eating habits are showing a diversified development trend.Compared with pork,lamb contains less fat and cholesterol,higher protein content and more delicious meat,which is becoming more and more popular among consumers.With the increasing consumption of mutton by residents,the gap between supply and demand of mutton in China has increased year by year.From 2010 to 2020,the mutton gap has soared from 40,000 tons to nearly 400,000 tons.China’s mutton sheep market mainly depends on the supply of local sheep varieties,but compared with foreign excellent meat breeds,local sheep have the disadvantages of slow growth and small carcass.Therefore,improving the meat production performance of local sheep is the most fundamental way to make up for the mutton supply gap.In this study,533 Grassland short tail sheep were selected as the research objects,and the polymorphic loci of MYF5 gene were screened and associated with body weight,body size,eye muscle area and backfat thickness were analyzed,so as to explore the genetic loci related to growth and meat production performance of Grassland short tail sheep,providing a theoretical basis for improving the performance of Grassland short tailed sheep.1.Association analysis of MYF5 gene polymorphism with growth and carcass traits in Grassland short tail sheepIn 533 Grassland short tail sheep populations,7 SNPs were screened by MYF5 gene,of which 4 SNPs were reported for the first time.The results of polymorphism analysis showed that SNP2,SNP3 and SNP4 were in moderate polymorphism(0.25<pic<0.5),and the other loci were in low polymorphism(PIC<0.25).All 7 SNPs are in Hardy Weinberg equilibrium.For the SNP1,the animals with AA genotype had significantly higher BW,WH,CC,and CBC than GA and GG ones(p<0.05).And the CW of AA genotype individuals were significantly higher than that of GG genotype individuals(p<0.05).The body weight of individuals with GG genotype of SNP2 was significantly higher than that of individuals with GT and TT genotype(P<0.05).For SNP3,the EMA of AC genotype individuals was significantly higher than that of CC genotype(p<0.05).For SNP4,The heterozygous genotype CT had more advantages in BW,BL,WH,CC,CBC and EMA than CC genotypes(p<0.05).Meanwhile,TT genotype also had higher mean values of BW,CBC,and EMA than CC genotype.About indicators WH and CC,CT genotype were significantly higher than CC and TT genotypes(p<0.05),and there was no significant difference between CC and TT genotypes(p>0.05).For SNP5,The CW of GG genotype was significantly higher than that of AA genotype,and there was no significant difference between GA genotype and the other two genotypes.For SNP6.the individuals with GG genotype had significantly higher values in BW.WH and CBC than individuals with TT genotype(p<0.05).Genotype GT was moderate in all indicators,except CBC was significantly lower than GG genotype,that had no significant difference(p>0.05)with the other two genotypes.For SNP7,the animals with TT genotype had higher BW and greater WH than CT and CC genotype(p<0.05).Besides,TT genotype had significant higher mean values of chest circumference(CC)than CC genotype,and there was no difference(p>0.05)between CT and CC genotype.Linkage disequilibrium analysis showed that SNP4 and SNP5(d’=0.97.R2=0.73),SNP6 and SNP7(d’=0.82,R2=0.64),SNP1 and SNP2(d’=0.98,R2=0.22)were strongly linked,and the linkage relationship ranged from strong to weak.In Grassland short tail sheep,seven SNPs could form four haplotype modules.The results of haplotype and haplotype combination analysis showed that the frequency of 8 haplotypes in 533 Grassland short tail sheep population was more than 1%,and the frequency of 4 haplotype combination was more than 5%.The correlation analysis between haplotype combination and phenotype showed that H1H3 and H2H3 haplotype combination showed great advantages,and the weight and body size traits were significantly greater than those of HI H1 and H1H2 haplotype combination.We also found that individuals with H3 haplotype have better growth performance,so it is speculated that H3 may be an advantage haplotype.Therefore,all individuals are divided into H3 containing group(H3+)and non H3 containing group(H3-).The results show that individuals with H3 haplotype have better growth performance,H3 haplotype is the advantage haplotype related to growth performance in Grassland short tail sheep.2.Effect of SNP in promoter region of MYF5 gene on gene transcriptional activityAmong the seven SNPs identified,three SNPs(SNP1-3)were located in the promoter region of MYF5 gene and were closely related to phenotypic data.In order to verify whether the three SNPs affect the changes of related phenotypes by affecting their promoter activity,firstly,the double luciferase reporting technology was used to determine that the core region of MYF5 gene promoter of Grassland short tail sheep is 5]4bp～241 bp upstream of the starting codon.Although SNP1,2 and 3 are not in the core region of their promoter,their region also has certain activity,and the online prediction software is used(AliBaba2.1,http://generegulation.com/pub/programs/alibaba2/index.html)It shows that SNP2 is in the binding region of transcription factor SP1(specific protein 1).The wild-type and mutant sequences of SNP2 were constructed by site directed mutation technology.Through the analysis of double luciferase reporting system,it was found that the double luciferase activity of mutant SNP2 was significantly lower than that of wild-type(P<0.01),and after exogenous addition of SP1 inhibitor,it was found that the double luciferase activity in this region also decreased significantly(P<0.05),Based on the above results,SNP2 may affect the transcription efficiency of MYF5 gene by changing the binding efficiency of transcription factor SP1 and its target sequence,so as to affect the related phenotypes of Grassland short tail sheep to a certain extent.The specific regulatory mechanism needs to be verified by more rigorous molecular biology experiments. |
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