香猪MAP3K4基因结构变异多态性和基因表达研究

Study of the Polymorphisms of Structural Variation and the Expression of MAP 3K4 gene in Xiang Pig

基于前期比较基因组学分析,在香猪的丝裂原活化蛋白激酶激酶激酶4(mitogen-activated protein kinase kinase kinase 4,MAP 3K4)基因第15内含子中发现一个189 bp的结构变异(MAP3K4-I15-sv189)。为探究该结构变异(structural variation,SV)在香猪群体中的多态性分布及其对基因表达的影响,本研究设计一对特异性引物,建立MAP 3K4基因SV的PCR检测技术,分析香猪群体中该结构变异的基因类型;采用RT-qPCR研究结构变异不同基因型之间基因的差异表达。结果显示,香猪群体中MAP3K4-I15-sv189位点存在两种基因型,DD型和ID型(D为缺失基因,I与参考基因一致),其中D等位基因的频率为97.35%;大白猪群体中检测到3种基因型,DD型、ID型和II型,D等位基因频率为80%。相比之下,香猪的D等位基因频率显著高于大白猪的相应值(P<0.05)。采用生物信息学方法分析结构变异区域得到多种剪接元件和重复元件,可能影响MAP 3K4基因的表达。比较香猪各组织中MAP 3K4基因的表达量,发现该基因在肺组织的表达量最高。进一步检测肺组织中3种结构变异类型的相对表达量,显示:DD型>ID型>II型,表明SV的缺失可能促进MAP 3K4基因的表达。本研究表明,香猪MAP 3K4的结构变异MAP3K4-I15-sv189以缺失型为主,缺失型可促进肺组织MAP 3K4基因的表达。研究结果有助于解析香猪易感肺部疾病的分子机制。

Based on previous data from comparative genomics,a structural variation(SV)with 189 bp in length was identified from the intron 15 of mitogen-activated protein kinase kinase kinase 4(MAP 3K4)gene in Xiang pig,which was defined as MAP3K4-I15-sv189.To explore the SV polymorphism in pig populations and its effects on gene expression,a PCR method was established using specific primers to discriminate SV genotypes and the gene expression level was determined via RT-qPCR technology.There found two genotypes,DD and ID,at the MAP3K4-I15-sv189 locus in Xiang pig population,in which the gene without SV region was indicated as D,while the reference gene containing the SV region was termed to be gene I.Of those,the frequency was 97.35%of allele D in Xiang pig group population.In Yorkshire pig population,three genotypes,DD,ID and II,were measured with frequency of 80%in allele D.And frequency of allele D in Xiang pigs was significantly higher than that in Yorkshire pigs(P<0.05).The elements in the SV region were analyzed via Bioinformatics methods.The deletion of SV might lead to the loss of several repeat elements,which might change the expression of MAP 3K4.A higher expression level of MAP 3K4 mRNA was then tested from the lung of Xiang pigs compared with other tissues.And the highest level of MAP 3K4 was found in the lung with genotype DD and the lowest from samples with genotype II.It indicated that the deletion of SV MAP3K4-I15-sv189 could accelerate the expression of MAP 3K4 in pig lung.The results illustrated that the SV MAP3K4-I15-sv189 in MAP 3K4 gene was dominated with genotype of deletion which resulted in the increase of gene expression in the lung,and would benefit to understand better undestanding the mechanism of susceptibility to diseases in Xiang pig lung.