犬MC1R基因多态性的研究

Research on the Polymorphism of MC1R Gene in Dogs

目的 利用PCR RFLP和PCR SSCP技术分别检测犬MC1R基因中两个序列T10 5A和R30 6Ter的基因多态性 ,为遗传育种、培育出更加优良的实验用犬奠定基础。方法 以 2 2 4只犬为实验材料 ,对犬MC1R基因T10 5A片段和R30 6Ter片段分别进行PCR RFLP和PCR SSCP分析 ,并且分别对具有RFLP和SSCP多态性的片段进行克隆测序 ,找出等位基因的差异位点。结果 经对犬MC1R基因的PCR RFLP分析 ,发现犬MC1R基因的T10 5A序列具有多态性 ,表现为三种基因型 :AA、AB和BB。通过对具有RFLP多态性的片段进行克隆测序发现MC1R基因在编码第 10 5位氨基酸的密码子处存在由G到A的单碱基突变 ,该突变导致第 10 5位氨基酸发生由精氨酸向苏氨酸的改变 ,此外在第 2 0 7位氨基酸的密码子处还发现由A到G的单碱基突变 ,并产生了一个HhaⅠ限制性内切酶位点。同时 ,经过犬MC1R基因的PCR SSCP分析 ,发现犬MC1R基因的R30 6Ter序列具有多态性 ,表现为三种基因型 :CC、CD、DD。通过对具有SSCP多态性的片段进行了克隆测序发现 ,MC1R基因在编码第 30 6位氨基酸的密码子处存在一个由CGA到TGA的终止突变 ,而使翻译终止。结论 利用PCR RFLP和PCR SSCP分析技术证实犬MC1R基因具有明显的多态性。

Objective The genetic polymorphism of two sequences T105A and R306Ter in the canine MC1R was detected by the way of PCR-RFLP and PCR-SSCP in order to serve for genetic breeding and cultivating better experimental canine.Methods The analysis of two sequences T105A and R306Ter in the canine MC1R gene was performed by the way of PCR-RFLP and PCR-SSCP in 224 canines,moreover PCR fragments with the polymorphism of PCR-FRLP and PCR-SSCP were sequenced respectively.Results By the analysis of PCR-RFLP,the T105A polymorphism was found with three genotypesAA,AB and BB.According to the result of sequencing one base change from G to A at the position 105 was found in the sequence of T105A,altering amino acid at the position 105 from arginine to threonine,furthermore another base change from A to G introduced a new HhaⅠ cut site at the position 207.At the same time,the R306Ter polymorphism was found with three genotypes CC,CD,DD by the analysis of PCR-SSCP.According to the result of sequencing one terminal change from CGA to TGA at the position 306 was found in the sequence of R306Ter and it made the termination of translation. Conclusions It can be proved by the analysis of PCR-RFLP and PCR-SSCP that the MC1R gene has significant polymorphism.