麦洼牦牛TLR1～10基因克隆及分子生物学特征分析  

Molecular Cloning and Biological Characterization Analysis of Toll-like Receptors 1-10 in Maiwa Yak

作　　者：林宝山[1,2] 兰道亮[1,2] 陈亚冰[1,2] 黄偲[1,2] 符梅[1,2] 李解[1,2] 李键[1,2] 

机构地区：[1]西南民族大学生命科学与技术学院 [2]西南民族大学青藏高原研究院,成都610041

出　　处：《畜牧兽医学报》2015年第5期728-737,共10页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：西南民族大学中央高校青年教师基金项目(2014NZYQN37);西南民族大学研究生学位点建设项目(2014XWD-S0906)

摘　　要：本研究克隆了麦洼牦牛天然免疫受体1~10基因的编码区,利用生物信息学工具分析基因特点,荧光定量PCR测定TLRs基因在不同组织中的表达量。序列比较分析结果表明,麦洼牦牛TLR基因与其他物种在核苷酸水平及氨基酸水平上均表现出很高的保守性。遗传进化方面,麦洼牦牛TLRs与牛和绵羊TLRs遗传进化距离最近,并与人、马、鼠TLRs等形成哺乳动物的一个分支,与鸡则形成遗传距离较远的一个分支。同时我们在进行系统发育分析时发现,TLR1、TLR6先聚为一小支,再与TLR10又聚为更紧密的一支,然后TLR1、2、6、10和TLR7、8、9分别聚集在两个单个的分支上,TLR其他成员各自成为一支。荧光定量结果表明,TLRs在麦洼牦牛各组织均有表达,但不同成员在不同组织的表达存在较大的差异。其中TLR2、TLR4和TLR6在脾表达量最高,在卵巢、小肠、肾、肝中有高表达,TLR1、TLR5、TLR7、TLR8、TLR9和TLR10在肾表达量最高,在肝、肾、脾等组织中高表达。综上所述,本研究的开展能为以后揭示TLRs在牦牛等高原模式动物分子免疫机制以及牦牛抗病育种奠定基础。In this study,the coding region of Maiwa yak toll-like receptors 1-10 genes were cloned,and then the bioinformatics tool was used to analyze the characteristics of TLRs sequences,and real-time PCR was used to detect the expression pattern of TLRs.The results showed that Maiwa yak TLRgenes exhibited high homologies at the nucleotide and the amino acid levels with other species.Phylogenetic relationships showed that Maiwa yak had a nearest relationship with cattle and sheep,Maiwa yak was clustered into a branch with human,horse and mouse.It was noteworthy that TLR1,TLR6 first clustered into a small branch,then gathered with TLR10 for a closer one,then TLR1,2,6,10 and TLR7,8,9were gathered in the 2individual branches,respectively.The other members of TLRs became respective one.Real-time PCR results showed that TLRs were expressed in all tissues of yak,but different members had different expression patterns.TLR2,TLR4 and TLR6had the highest expression in the spleen,followed with ovary,small intestine,kidney,liver.TLR1,TLR5,TLR7,TLR8,TLR9 and TLR10had the highest expression in kidney,higher expression in liver,kidney,spleen.The study provide useful information for further study in immune molecular mechanisms and disease resistance breeding of yaks.

关 键 词：麦洼牦牛 TLR1-10 克隆 荧光定量PCR 组织表达分布 

分 类 号：S823.85[农业科学—畜牧学]