孕酮受体基因外显子4作为小尾寒羊多胎候选基因的研究

Study of PGR-exon 4 as a Candidate Gene for Fecundity in Small Tail Han Sheep

试验采用53只小尾寒羊母羊(山东梁山和泰安)、25只无角陶赛特羊母羊(甘肃永昌肉用种羊场)、35只蒙古羊母羊(甘肃武威)。分别采集颈静脉血样提取DNA,利用GenBank中人孕酮受体基因外显子4和绵羊孕酮受体基因mRNA序列,设计绵羊孕酮受体基因外显子4限制性片段特异性引物,进行PCR扩增,对其PCR产物克隆、测序验证。用PCR-SSCP检测多态性,通过DNA测序确定单核苷酸多态位点(SNPs)。经过研究分析表明,小尾寒羊由野生型AA型、杂合AB型、突变纯合BB型3种基因型组成,蒙古羊由AA型、AB型两种基因型组成,无角陶赛特羊只有AA型一种基因型,不存在突变;在基因型分布上小尾寒羊与无角陶赛特羊和蒙古羊之间存在极显著差异(P<0.01),而无角陶赛特羊和蒙古羊之间差异不显著(P>0.05);小尾寒羊突变纯合基因型BB和杂合AB中存在4个突变位点,此外在发现的4个突变位点中,只有第4个突变位点246碱基T→C引起氨基酸突变。

The DNA samples were purified from the blood samples taken from 53 Small Tail Han sheep ewes in Shandong and 25 Poll Dorset ewes from Yongchang of Gansu, 35 Mongolia ewes from Wuwei of Gansu. The specific constriction primer of sheep was constructed by using GenBank human PGR-exon 4 and sheep PGR mRNA sequence. The PCR amplification was done and the cloning of its products was made subsequently for the analysis. The PCR-SSCP technology was used for Polymor- phism analysis and DNA sequence analysis technology was used for SNPs analysis. The results showed in the following：There were three genotypes of AA wild type, AB heterozygote and BB mutative homozygote in Small Tail Han sheep, there were only two genotypes of AA and AB in Mongolia sheep, there was only AA genotype found in Poll dorset and there are no other genotypes has been found; The statistic analysis show that the genotype constitution of Small Tail Han sheep showed markedly difference to poll Dorset and Mongolia sheep （P〈0.01） while there were no significance difference between Poll Dorset and Mongolia sheep（P〉0.05） ;There were 4 mutative loci in the genotype of mutative homozygote BB and heterozygote AB of the Small Tail Han sheep, In addition, only fourth mutative loci of 246 T→C leads the mutation of amino acids.