Analysis Of MC4R, POMC, CSN3and BLG Gene Polymorphisms And Their Association With Growth Traits In Hu Sheep And East Friesian Crossbred Sheep

A total of298individuals were sampled from two sheep populations:Hu sheep （n=168） and East Friesian×Hu crossbred sheep （F1, n=130）. Based on the analysis of production performance, the DNA pool PCR amplified products sequencing technique was used to detect the polymorphism of MC4R, POMC, CSN3, β-lg genes, and34SNPs were detected, PCR-RFLP and CRS-PCR-RFLP techniques were applied to detect the genetic structure of7SNPs in the MC4R, POMC, CSN3and β-lg genes in these populations to understand the molecular genetic information concerned candidate gene loci. Association analysis were carried out to evaluate the effects of genotypes of candidate genes on production traits of the two sheep breeds to look for the molecular marker or QTL of economic traits, therefore to provide scientific information for breeding of dual purpose （milk and meat） sheep. The results were as follows:1、Production performance analysis of Hu sheep and East Friesian×Hu crossbred sheepHu sheep and East Friesian×Hu crossbred sheep aged six months （n=10） were used to evaluate the meat performance, and the20th day after delivery of Hu sheep and East Friesian x Hu crossbred sheep （n=10） were used to evaluate the milk performance. The results showed that the net meat rate of Hu sheep was significantly higher than that of East Friesian crossbred sheep（P<0.05）; the antemortem live weight, carcass weight, slaughter rate, meat-bone ratio, fleshcolor, pH value, water lose rate and eye muscle area of Hu sheep were higher than that of East Friesian x Hu crossbred sheep（P>0.05）. The analysis of milking performance showed that the concentrations of lactose, milk protein and milk fat percentage in the two groups were significantly different （P<0.01）, the milk fat percentage of East Friesian×Hu crossbred sheep was significantly higher than that of Hu sheep, but the concentrations of lactose and milk protein were opposite.2、Polymorphism of MC4R gene and its association with growth traits in Hu sheep and East Friesian×Hu crossbred sheepThe DNA pool PCR amplified products sequencing technique was applied to detect the polymorphism of MC4R gene and10mutations （NM₀01126370）: g.110C/G, g.222C/T, g.306G/A, g.548C/T, g.706C/A, g.732C/G, g.951G/A, g.1229G/A, g.1267G/A and g.1291G/C were detected. The g.110C/G were in the5’UTR region, g.222C/T, g.306G/A, g.548C/T, g.706C/A, g.732C/G and g.951G/A were in the coding region, and g.1229G/A, g.1267G/A and g.1291G/C were in the3’UTR region. The mutation of548and732sites were missense mutations which caused changes of Thr to Met and Ile to Met, respectively. The PCR-RFLP technique was applied to detect the genetic structure of loci306and1267, and the CRS-PCR-RFLP technique was applied to detect the genetic structure of loci548and706. The genetic diversity analysis revealed that the locus306was moderate polymorphic locus, the loci548,706and1267were low polymorphic loci of the two sheep breeds. The population at all of the four SNPs was at Hardy-Weinberg unbalance （P<0.05）.Association analysis of the four loci with the body weight demonstrated that in the two sheep breeds there was no significant association between different genotypes at locus306in the body weight （P>0.05）. In the Hu sheep, CT genotype at locus548significantly differ from CC genotype in the six months weight and4-6months average daily gain （P<0.05）. The six months weight and4-6months average daily gain of CA genotype at locus706were significantly higher than that of CC genotype （P<0.01）. The two months weaning weight and four months weight of GA genotype at locus1267were significantly higher than that of GG genotype （P<0.01）, and the six months weight of GA genotype was significantly higher than that of GG genotype （P<0.05）. In East Friesian×Hu crossbred sheep, the genotypic at locus548affected the four months weight and six months weight highly significantly （P<0.01）, and affected the4-6months average daily gain significantly （P<0.05）. The birth weight, two months weaning weight, six months weight and4-6months average daily gain of AA genotype at locus706were significantly higher than that of CC genotype （P<0.01）. The genotypic at locus1267affected four months weight and six months weight at0.05and0.01level, respectively.Association analysis of the four loci with four months body sizes showed that in Hu sheep, the hip width of AA genotype at locus306was higher than that of CA genotype （P<0.05）; at locus548the body length of CT genotype was significantly higher than that of CC genotype （P<0.01）, the chest circumference of TT genotype was significantly higher than that of CC and CT genotype （P<0.01）; at locus706the body length of CA genotype was significantly higher than that of CC genotype （P<0.05）; at locus1267the body height and rump height of GA genotype were significantly higher than that of GG genotype （P<0.05）. In East Friesian×Hu crossbred sheep, the cannon circumference of GG and GA genotype at locus306were significantly higher than that of AA genotype （P<0.01）; the body height （P<0.01） and rump height （P<0.05） of CT genotype at locus548were significantly higher than that of CC genotype; at locus706, the body height, body length, hip width and cannon circumference of AA genotype were significantly higher than that of CC genotype （P<0.01）, the chest depth of AA genotype was significantly higher than that of CC genotype （P<0.05）; at locus1267the body height and cannon circumference of GA genotype were significantly higher than that of AA genotype （P<0.05）.3、 Polymorphism of POMC gene and its association with growth traits in Hu sheep and East Friesian×Hu crossbred sheepThe DNA pool PCR amplified products sequencing technique was applied to detect the polymorphism of POMC gene and two silent cSNPs mutations （NM₀01009266.1）:g.273T/C and g.456G/A in the exon3of POMC gene were identified. The PCR-RFLP technique was applied to detect the genetic structure at locus273, and the genetic diversity analysis showed that locus273was moderate polymorphic locus in Hu sheep, while the CC genotype was not detected in East Friesian×Hu crossbred sheep and was low polymorphic locus. The population at g.273T/C was at Hardy-Weinberg unbalance （P<0.05）.Association analysis of locus273with body weight and four months body sizes showed that in Hu sheep the two months weaning weight, four months rump height of CC genotype and the four months body length, cannon circumference of TC genotype were significantly higher than that of TT genotype （P<0.05）; the four months weight and six months weight of CC genotype were significantly higher than that of TT and TC genotype （P<0.01）; the four months body height （P<0.01） and body length （P<0.05） of CC genotype were significantly higher than that of TT and TC genotype; the four months cannon circumference of CC genotype was significantly higher than that of TT genotype （P<0.01）. In East Friesian×Hu crossbred sheep the two months weaning weight, four months weight, body height, body length, chest depth and cannon circumference of TC genotype were significantly higher than that of TT genotype （P<0.05）; and the six months weight of TC genotype was significantly higher than that of CC genotype （P<0.01）.4、 Polymorphism of CSN3and β-lg genes and its association with growth traits in Hu sheep and East Friesian x Hu crossbred sheepThe DNA pool PCR amplified products sequencing technique was applied to detect the polymorphism of CSN3and β-lg gene, one SNP mutation in exon4of CSN3gene （NM₀01009378.1）:g.237T/C and21SNPs in β-lg promoter region （FR821261.1）:g.167A/G, g.213T/C, g.235C/T, g.237C/T, g.239T/G, g.333G/T, g.357T/C, g.368T/G, g.378C/T, g.415A/G, g.517C/A, g.700G/C, g.711G/A, g.1384G/A, g.1573G/A, g.1603C/T, g.1620A/G, g.1652C/G, g.1671T/C, g.1701A/C and g.1902A/T were detected. The PCR-RFLP and CRS-PCR-RFLP technique were used to detect genetic structure at locus237of CSN3gene and locus1652of β-lg gene, respectively. The genetic diversity analysis showed that locus237was low and moderate polymorphic locus in Hu sheep and East Friesian×Hu crossbred sheep, respectively; locus1652was low polymorphic locus in both of the two breeds. The population at both of the two loci was at Hardy-Weinberg unbalance （P<0.05）.Association analysis of locus237with body weight and four months body sizes showed that there was no obvious difference with growth traits in both of the two breeds （P>0.05）.Association analysis of locus1652with body weight and four months body sizes showed that in Hu sheep the four months weight and body height of CC genotype were significantly higher than that of GG genotype （P<0.01）; the four months chest circumference, Hip width and Cannon circumference of CC and GG genotype were significantly higher than that CG genotype （P<0.05）. In East Friesian×Hu crossbred sheep the six months weight and four months body height of GG genotype were significantly higher than that of CC genotype （P<0.05）； the four months chest circumference and rump height of GG genotype were significantly higher than that of CG genotype （P<0.05）.