牦牛TLR7基因的克隆序列分析及其组织表达研究  

Cloning and Sequence Analysis of TLR7 Gene and Its Expression and Distribution in Tissues of Yak

作　　者：林宝山[1] 兰道亮[1] 陈亚冰[1] 黄偲[1] 梁彦彦[1] 李键[1] 

机构地区：[1]西南民族大学生命科学与技术学院,青藏高原研究院,四川成都610041

出　　处：《中国畜牧杂志》2014年第19期10-16,共7页Chinese Journal of Animal Science

基　　金：中央高校青年教师基金项目(2014NZYQN37)

摘　　要：为了解牦牛TLR7基因特点及其在牦牛各组织器官动态表达分布规律。根据GenBank中牛TLR7序列设计引物克隆并对所得序列进行生物信息学分析,并以β-actin为参照基因,SYBR Green I为荧光染料,建立检测牦牛天然免疫受体TLRs家族TLR7基因的荧光定量PCR方法,对TLR7基因在牦牛各组织中的表达分布进行研究。结果表明:克隆得到牦牛TLR7基因编码区全长3 153 bp,编码氨基酸1 050个,N端含有27个氨基酸组成的信号肽,分子量120.7192 ku,理论等电点为7.35;TLR7编码蛋白整体表现为亲水性;10条序列当中牦牛与野牦牛、普通牛、绵羊等物种具有较高的同源性,在系统发育树中距离最近,说明TLR7基因序列具有较高的保守性;荧光定量分析发现,TLR7基因在牦牛所有组织中均有表达,TLR7基因在肾脏表达量最高,在大肠表达量最低,在肝、脾、肺、卵巢、小肠、心、胃、肌肉、乳腺等组织依次有较高的表达。该研究结果为理解TLRs在牦牛等其他高原动物体内的分布及动态表达规律以及牦牛抗病毒免疫及育种的相关研究奠定了基础。To understand the characteristics and the dynamic distribution of yak TLR7 gene in various tissues.According to Bovine TLR7 sequence in GeneBank, we designed cloning primers and gain the sequence of TLR7 gene and sequentially perform related bioinformatic analysis. In addition, the study also successfully established a real-time PCR methods for detecting the expression of TLR7 gene in yak, β-actin as chosen a reference gene. The results of sequence analysis showed that the length of the cloning TLR7 cDNA is 3 153 bp. It encodes a peptide of 1 050 amino acids whose calculated molecular weight is 120.7192 ku, PI is 7.35. The predicted results also showed that TLR7 protein perform the hydrophilic. Sequence homology analysis showed that yak has higher homology with Wild Yak,cattle, sheep and other species, which have the nearest perimeter in the phylogenetic tree. The results of real-time PCR showed that TLR7 gene was expressed in all tissues of yak, highest in the kidney and lowest in the large intestine. Moreover, it has relatively higher expression in the liver, spleen, lung, ovary, small intestine and other tissues. The results suggested that TLR7 gene may play an important role in the molecular mechanisms of yak immune disease, which make a foundation for understanding the dynamics expression pattern of the of TLRs in yaks and other animals in the highland and provides a basis for future research in disease resistance breeding of yak.

关 键 词：TLR7 克隆 荧光定量PCR 组织表达分布 牦牛 

分 类 号：S823.2[农业科学—畜牧学]