哈萨克绵羊DRB1基因外显子1和4多态性与布鲁氏菌病的相关性研究

The Correlation Between Polymorphisms of Exon1 and Exon4 of DRB1 Gene and Brucellosis in Kazakh Sheep

试验旨在对哈萨克绵羊DRB1基因外显子1和4多态性与布鲁氏菌病的相关性进行研究。使用虎红平板凝集试验(RBPT)对试羊的血清进行血清学检测,参考GenBank中绵羊MHC ClassⅡ区DRB1基因序列(登录号:NC_040271.1),对其外显子1和4片段设计引物,采用PCR-SSCP和DNA测序技术对230只哈萨克绵羊的DRB1基因进行多态性检测,分析其多态位点与哈萨克绵羊布鲁氏菌易感性之间的关系。RBPT检测发现66只哈萨克绵羊为布鲁氏菌感染阳性,阳性检出率为28.7%。外显子1片段存在一个SNP位点(F1-G22A),测序确定两种基因型(GG、GA),优势等位基因和基因型分别为G、GG,F1-G22A多态位点的易感基因型为GA。卡方检验表明,哈萨克绵羊DRB1基因F1-G22A多态位点与布鲁氏菌易感性的相关性不显著(P>0.05)。通过生物信息学在线软件分析得出,F1-G22A多态位点导致了RNA二级结构的改变和最小自由能的降低,引起了蛋白质二级结构的改变。DRB1基因外显子4片段未发现SNPs。由此得出,哈萨克绵羊DRB1基因F1-G22A多态位点与布鲁氏菌易感性可能存在一定的相关性。

The purpose of this experiment was to study the correlation between exon 1 and 4 polymorphisms of DRB1 gene and brucellosis in Kazakh sheep.Using RBPT serological tests to try sheep serum,reference in GenBank sheep MHC ClassⅡarea DRB1 gene sequences(Accession No.:NC_040271.1),the exon 1 and 4 pieces designed primers,using PCR-SSCP and DNA sequencing technology to 230 Kazak sheep DRB1 gene polymorphism detection,analyze its polymorphism loci and the relationship between the Kazak sheep Brucella susceptibility.The results showed that 66 Kazakh sheep were positive for Brucella in RBPT test,and the positive detection rate was 28.7%.There was one SNP locus(F1-G22A)in exon 1 fragment,and sequencing determined two genotypes(GG and GA),the dominant allele and genotype were G and GG respectively,and the susceptibility genotype of the polymorphisms of F1-G22A was GA.Chi-square test showed that there was no significant correlation between the polymorphisms of DRB1 gene F1-G22A and Brucella susceptibility in Kazakh sheep(P>0.05).According to the analysis of bioinformatics online software,the F1-G22A polymorphic sites lead to the change of RNA secondary structure and the decrease of minimum free energy,and lead to the change of protein secondary structure.No SNPs were found in DRB1 exon 4 fragment.Therefore,there might be a certain correlation between the polymorphisms of DRB1 gene F1-G22A and Brucella susceptibility in Kazakh sheep.