保山猪和杜×保仔猪MC1R基因型鉴定与分析

Characterization and analysis of MC1R genotype in Baoshan pig and Du×Bao hybrid pig

为了解释保山猪和杜洛克猪杂交后代出现毛色分离的原因,寻找判定保山猪纯种与否的分子标记,试验采用PCR扩增、分子克隆与多重序列比对技术,对保山猪MC1R基因编码区序列及杜×保仔猪(S1~S6窝)MC1R基因型进行分析。结果表明:保山猪MC1R基因编码区序列长度为963 bp,共编码321个氨基酸。以野猪MC1R基因编码区序列为参考,保山猪存在5个SNP位点,在翻译水平上发生2个错义突变及3个同义突变,2个错义突变为第283位碱基由G>A、第305位碱基由T>C,3个同义突变为第51位碱基由G>A,第363位碱基由T>C,第729位碱基由G>A,证实调控保山猪黑毛色的等位基因为E^(D1)。S1~S4窝为纯种的杜×保仔猪,仔猪均为全黑色,在碱基491C>T、727G>A、729C>A处均存在突变,这3个SNP位点处图谱均为双峰,基因型为E^(D1)/e;S5窝为杂种的杜×保仔猪,为黑毛、黄毛仔猪,黑毛仔猪基因型为E^(D1)/e,黄毛仔猪基因型为e/e;S6窝为杂种的杜×保仔猪,为黑毛及黑黄纵纹仔猪,其基因型均为E^(D1)/e。说明MC1R基因可以作为解释保山猪毛色分离和判断纯种与否的分子标记。

In order to explain the reason of coat color separation in thehybrid of Duroc and Baoshan pig and seek molecular marker to determine the purity of Baoshan pig.PCR amplification,molecular cloning and multiple sequence alignment techniques were used to analyze MC1 R gene CDS in Baoshan pig and MC1 R genotype in its hybrid piglets(S1-S6 litters).The results showed that the MC1 R gene CDS of Baoshan pig was 963 bp in length,encoding 321 amino acids.Compared with wild boar,Baoshan pig had five SNPs in the coding region of MC1 R gene.Two missense mutations and three synonymous mutations occur at the translation level.The two missense mutations were the 283 st(G>A),the 305 st(T>C),and the three synonymous mutations were the 51 st(G>A),the 363 st(T>C),the 729 st(G>A)bases.This indicated that the allele regulating the black hair color of Baoshan pig was E^(D1).The S1~S4 litters of piglets were the purebred Du x Bao pigs,and they were all black,which had three SNPs at base 491 C>T,727 G>A and 729 C>A,and the DNA profiles showed that this three SNPs are all double peaks,whose genotype was E^(D1)/e.The S5 litter of piglets were the hybrid Du x Bao pigs,and their offspring appeared black piglets and yellow piglets.The genotype of black piglets and yellow piglets were E^(D1)/e,e/e.The S6 litter of piglets were hybrid Du x Bao pigs,and their offspring appeared black piglets and black-yellow striped piglets and the genotype was E^(D1)/e.It is suggested that MC1 R gene can be used as a molecular marker to explain the color separation of Baoshan pig and determine whether it is a pure breed or not.