猪SLA-DRA基因SNP位点筛选及其生物信息学分析

SNP screening and bioinformatics analysis of SLA-DRA gene in pigs

以八眉猪、蕨麻猪和甘肃黑猪为研究对象,利用PCR-SSCP和测序方法对SLA-DRA基因4个外显子进行SNP位点筛选,并利用生物信息学软件预测各SNP位点对DRA基因mRNA二级结构、蛋白质二级结构和三级结构的影响。结果在3个猪种中共检测到6个DRA基因SNP位点,分别为外显子1的A^(177)G,外显子2的A^(3093)C,以及外显子4的A^(4167)G、A^(4246)G、C^(4282)T和G^(4293)A,其中A^(3093)C、A^(4167)G和G^(4293)A为错义突变,分别导致甲硫氨酸(Met)变为亮氨酸(Leu)、谷氨酰胺(Gln)变为精氨酸(Arg)和精氨酸(Arg)变为组氨酸(His)。以3个错义SNP位点为基础,构建了6种DRA单倍型,其中H1、H3和H6为主要单倍型,频率分别为0.25、0.21和0.21。生物信息学分析表明,三个猪种DRA基因单倍型的mRNA二级结构、蛋白质二级结构和三级结构与野生型均存在一定的差异。研究结果可为八眉猪、蕨麻猪和甘肃黑猪的保种和抗病育种提供理论依据。

The single nucleotide polymorphism （SNP） site of four exons of SLA-DRA gene in Bamei, Juema and Gansu Black pigs were screened via polymerase chain reaction-single strand conformation polymorphism （ PCR-SS- CP） and DNA sequencing analysis, and further secondary structure of mRNA, secondary and tertiary structures of DRA protein were predicted through the bioinformatics tools. The results showed that 6 SNP sites were detected, in-cluding A177 G, located in exon 1, A3093 C, located in exon 2, A4167 G, A4246 G, C4282 T and Q4293 A , located in exon 4 , and among these SNP sites, A3093 C, A4167G and G4293A were missense mutation, A3093 C led to methionine （Met） into leucine （Leu） , A4167G led to glut amine （Gin） into arginine （ Arg）,and G4293 A led to arginine （ Arg） into histidine （His） . Based on the 3 missense SNPs, 6 haplotypes were constructed, in which HI, H3 and H6 were the main hap- lotypes with frequencies 0. 25, 0. 21 and 0. 21, respectively. The results of bioinformatics showed that secondary structure of mRNA, secondary and tertiary protein structures of DRA haplotypes in three pigs were different from those in wild type. The consequence would provide basis for study of breed conservation and molecular breeding in disease resistance for Bamei, Juema and Gansu Black pigs.