巴马香猪VRTN基因克隆、表达及其多态性与繁殖性状的关联分析

Cloning, Expression of VRTN Gene and Association Analysis of Its Polymorphism with Reproductive Traits in Bama Xiang Pigs

旨在研究VRTN(vertebrae development homolog)基因在巴马香猪群体中的编码区序列特征、组织表达情况、脊椎数性状因果突变位点ins291的等位基因频率及其与乳头数和产仔数性状的关联。本研究采集3头0日龄巴马香猪组织,利用cDNA克隆技术获得VRTN基因编码区全长序列并进行生物信息学分析,利用荧光定量PCR技术检测VRTN在心、肝、脾、肺、肾、背最长肌和皮下脂肪组织中的表达情况;采集279头经产巴马香猪母猪血液,检测ins291位点在该群体中的频率分布,并与乳头数、产仔数性状进行关联分析。结果表明,巴马香猪VRTN基因编码区全长2 097 bp,其编码的氨基酸序列在不同物种间存在大量保守区域;VRTN基因编码698个氨基酸,预测为亲水性蛋白质,存在1个螺旋转角螺旋域超家族结构功能区、2个低复杂度区域和2个内部重复结构,并与核受体辅抑制子1(NR6A1)、果蝇同源框基因Prospero的脊椎动物同源蛋白2(PROX2)和富亮氨酸重复序列74亚基(LRRC74A)等蛋白具有相互作用;VRTN基因在0日龄巴马香猪各组织中均有表达,其中在背最长肌组织中表达量最高;巴马香猪保种群体中VRTN基因有利突变位点ins291的等位基因频率为21.15%,不同基因型个体之间平均产仔数、总乳头数、左侧乳头数、右侧乳头数和单侧最大乳头数性状均无显著差异(P>0.05),但纯合突变型(ins/ins)个体的乳头数性状均高于其他基因型个体。结果提示,在巴马香猪产业化生产中可通过分子育种手段提高VRTN ins291有利等位基因频率,但不影响其产仔数性状。

The aim of this study was to investigate the sequence characteristics,tissue expressions of VRTN(vertebrae development homolog)gene in Bama Xiang pig population and the gene frequency of causal mutation site ins291 for vertebral number and its association with teat number and litter size.The tissues of three 0-day-old Bama Xiang pigs were collected,and the full-length sequence of VRTN gene coding region was obtained by cDNA cloning technology and bioinformatics analysis was also conducted.The expression of VRTN in heart,liver,spleen,lung,kidney,longissimus dorsi and subcutaneous fat tissue was detected by fluorescence quantitative PCR.Blood samples from 279 Bama Xiang sows were collected to test the frequency distribution of ins291 site in the population,and its association with teat number and litter size was analyzed.The result showed that the full-length of the VRTN gene coding region in Bama Xiang pig was 2097 bp,its amino acid sequence had lots of conserved regions among different species;VRTN gene encoded 698 amino acids,and it was predicted to express a hydrophilic protein.VRTN protein was most likely having one helix-turn-helix superfamily structural functional region,two low-complexity regions and two internal repetition structures.Moreover,the protein interacted with nuclear receptor subfamily 6 group A member 1(NR6A1),Prospero homeobox 2(PROX2)and leucine rich repeat containing 74A(LRRC74A).VRTN gene was expressed in all tissues of 0-day-old Bama Xiang pigs,with the highest expression in longissimus dorsi.The allele frequency of beneficial mutation site ins291 of VRTN in Bama Xiang pigs from the preserved population was 21.15%.There was no significant difference in average litter size,total teat number,left teat number,right teat number and maximum teat number of one side among individuals with different genotypes(P>0.05),but the teat number of individuals with homozygous mutant genotype(ins/ins)was higher than those with other genotypes.These results indicate that molecular breeding method can be used to increase the favorable allele frequency of VRTN ins291 in the industrialized productions of Bama Xiang pigs,which don’t affect litter size.