猪NLRC5基因克隆及序列特征分析

Cloning and sequence analysis of NLRC5 gene from pigs

为了解猪天然免疫受体NLRC5基因特征,用人类、小鼠等物种NLRC5基因已知片段设计分段扩增的引物,同时运用RACE技术,克隆该序列并进行序列生物信息学分析.基因序列分析表明,克隆得到猪NLRC5基因c DNA全长为6638bp,基因开放阅读框为5538bp,共编码1846个氨基酸.蛋白质预测结果显示,NLRC5编码蛋白质整体表现为亲水性,蛋白质结构域分析结果显示,NLRC5具有典型的NACHT区及20个LRRs.同源性分析表明猪与牛、绵羊、猕猴、人等物种具有较高同源性,达到了80%左右,遗传进化树分析表明与牛和绵羊进化关系最近.进一步组织表达分析表明NLRC5基因除在淋巴结、脾脏等系统免疫器官中具有较高表达量,在肠淋巴结、胃、肺等黏膜器官中也有较高的转录水平.本研究结果说明猪NLRC5基因在长期进化中较为保守,尤其在哺乳动物间具有较高的保守性,在免疫反应中可能扮演着重要的角色.猪NLRC5基因序列的成功克隆为后续NLRC5在机体内免疫系统中的功能研究奠定了理论基础.

In order to investigate the genetic characterization of NLRC5 in innate immune receptor of Sus scrofa, this research designed primers by using the known gene sequences of NLRC5 from human and mice. This research also applied RACE tech- neque to clone this sequence and conducted corresponding bioinformatics analysis. As shown in the analysis, the cloning process produced NLRC5 cDNA with a length of 6638bp,the open reading frame was 5538bp encoding 1846 deduced amino acids. The protein structure prediction showed that the protein deduced by NLRC5 was hydrophilic. The analysis of protein structural do- mains showed that NLRC5 had typical NACHT domain and 20 LRRs. The analysis of homology manifested that the gene of Sus scrofa was close to buffalo, Ov/s aries, Macaca mulatta and human, which is up to about 80%. The phylogenetic tree analysis proved that buffalo and Ov/s aries had the closest evolutionary relationship. Further analysis of tissues expression indicated that NLRC5 had higher transcriptional levd in tissues like intestinal lymphnode, stomach and lung, as weU as in immune organs hke lymphnodes and spleen. The research demonstrated that NLRC5 of Sus scrofa was stable in evolution, especially among mam- mals, and it might have played an important part in immune responses. The successful cloning of NLRC5 of Sus scrofa has estab- lished a theoretical basis for future research of NLRC5 function in immune system.