Association Between The Polymorphisms Of TYR、ASIP Gene And Coat Color In Sheep

Objective:In order to investigate the relationship between coat color of ovis and ASIP、TYRaffecting the mammaila coat.we analysised the correlation between sequence polymorphismin encoding region of ASIP、TYR and coat color in sheep by PCR-SSCP and detected therelative expression levels of mRNA (ASIP、TYR) from skin tissues of Chinese Merinosheep(Junken type)(white/Brown) and Altay sheep(Brown/Black).we analysised therelationship between the polymorphism in DNA/the change in mRNA and the coat color,itcan offer the basis for regulation mechanism proved coat color formation of sheep.Methods:1、Through the PCR-SSCP and the second generation sequencing technology,we screenedsome SNPs in the CDS region of ASIP and TYR and analysed the statistical correlationbetween SNPs which conform to SSCP electrophoresis rule and different wool color ofbreeds.2、We detected the relative expression quantity of ASIP and TYR gene in skin tissues ofChinese Merino sheep(Junken type) and altay sheep by the real-time fluorescence quantitativePCR.Result：1、Deletion and substitution mutations were detected in ASIP gene.5-bp deletion (D5：AGGAA) in exon2resulted in a frameshift mutation and the protein encoded by ASIP earlytermination of63amino acids. Substitution mutation T→A (g.5179bp) in4exon resulted toamino acid change (Cys→Ser). Two mutations were detected in Altay sheep,but one(5-bpdeletion in2exon) in Chinese Merino sheep(Junken type). The statistic results showed thatthere were no relationship between two SNPs and the coat color in Altay sheep（P>0.05）,butthe deletion mutations were extremely significant correlation with coat color of ChineseMerino sheep(Junken type)（P<0.01）.2、TYR gene were screened out four SNPs, respectively, the5’ end untranslated region G→C(g.683bp) substitution, exon2A→G (g.11139bp) transversion, intron3A→G (g.47998bp)transversion and exon5G→C (g.48446bp) substitution.All four SNPs were detected in Altaysheep,but two SNPs: intron3A→G (g.47998bp) transversion and exon5G→C (g.48446bp)substitution in Chinese Merino sheep(Junken type). g.683bp G→C mutation of TYR wasextremely significant correlation with coat color of Altay sheep（P<0.01）. The remainingmutations were not correlation with coat color.3、By the Q-PCR assay,we detected the relative expression level of ASIP、TYR,and the resultshowed that ASIP and TYR gene expressed in both altay sheep and Chinese Merinosheep(Junken type) skin tissues and there were no correlation between the relative expressionamount and the coat color.Conclusion：1、5-bp deletion (D5：AGGAA) in exon2of ASIP gene was detected in Altay sheep andthere was a significant correlation between the SNP and coat color of Chinese Merino sheep(Junken type)(P<0.01)，it can be used as marker gene related coat color in ChineseMerino sheep(Junken type). There was a significant correlation between the mutation g.683bpG→C of TYR gene,which can be used as marker gene related coat color in Altay sheep, andcoat color of Altay sheep (P<0.01).3、Both ASIP and TYR exprss in skin tissues of Chinese Merino sheep(Junken type) and Altaysheep,but the mRNA relative expression quantity of ASIP and TYR didn’t related to the coatcolor of Chinese Merino sheep(Junken type) and Altay sheep(P>0.05).