淇河鲫IL-8cDNA克隆及组织表达分析  

Cloning and tissue expression analysis of the interleukin 8 gene from crucian carp (Carassius auratus) in Qihe River

作　　者：王俊丽[1] 雒燕婷[1] 郝光[1] 孙君君[1] 段艳艳[2] 卢荣华[2] 李学军[2] 孔祥会[2] 聂国兴[2] 

机构地区：[1]河南师范大学生命科学学院,河南新乡453007 [2]河南师范大学水产学院,河南新乡453007

出　　处：《水产学报》2013年第9期1281-1289,共9页Journal of Fisheries of China

基　　金：国家自然科学基金项目(30972252);河南省重点科技攻关项目(122102110049)

摘　　要：IL-8是最早发现的趋化因子,属于CXC亚家族,与鱼类非特异性免疫水平关系密切。实验采用同源克隆和cDNA末端快速扩增(RACE)方法,从淇河鲫总RNA反转录产物中获得了641 bp IL-8cDNA序列,其中包含102 bp的5'非编码区,242 bp的3'非编码区和297 bp的开放阅读框。该基因编码由98个氨基酸残基组成的前体蛋白,前22个氨基酸残基为信号肽。肽链中含有4个保守半胱氨酸,前2个半胱氨酸被1个精氨酸分隔,形成CXC结构。第一个半胱氨酸前缺乏ELR基序,其组成是DPR。由氨基酸同源性分析和进化分析可知,淇河鲫IL-8与鲤、鳙、草鱼、鲢等鲤科鱼类的进化关系较近,与鲤IL-8的同源性最高(94%)。通过实时荧光定量PCR检测,淇河鲫IL-8在被检测的8个组织中都有表达,其中在肌肉中的表达量最高,其次为脾脏、头肾和脑等。研究结果对调控淇河鲫非特异性免疫能力,提高淇河鲫健康养殖水平具有一定参考价值。Interleukin 8（IL-8）,belonging to CXC family,is the first discovered chemokine,and is closely associated w ith non-specific immunity of fish.In this study,a full-length cDNA sequence of the IL-8 gene w as cloned from crucian carp（Carassius auratus） in Qihe River using RT-PCR and RACE techniques.The IL-8 cDNA sequence consisted of 641 bp nucleotides,including 102 bp at 5’UTR,242 bp at 3’UTR and an open reading frame w ith 297 bp nucleotides.This cDNA sequence encoded a precursor protein of 98 amino acids and the former 22 amino acid residues formed a signal peptide.The putative peptide contained four cysteine residues,and the former tw o cysteine residues w ere separated by an arginine residue and consequently formed a CXC structure.Furthermore,similar to the most fishes,IL-8 of crucian carp lacked the completed ELR（Glu-Leu-Arg） motif in front of the CXC structure,w hich w as commonly present in mammalian and chicken IL-8 molecules.Therefore,DPR motif w as substituted for ELR motif in IL-8 of crucian carp.Comparison of the amino acid sequences show ed that the crucian carp IL-8 shared the highest similarity w ith Cyprinus carpio（94%）.Phylogenetic analysis also demonstrated that the crucian carp IL-8 had the close relationship w ith Cyprinidae species such as Cyprinus carpio（94%）,Hypophthalmichthys nobilis（81%）,Ctenopharyngodon idella（81%） and Hypophthalmichthys molitrix（80%）.IL-8 mRNA expression in eight different tissues of crucian carp w as determined by Real-time Quantitative PCR technique.The results displayed that the level of IL-8 mRNA w as highest in muscle,follow ed by spleen,head kidney and brain.The research w ould be beneficial to the regulation of non-specific immunity in crucian carp,and also provides a certain reference value for improving the health breeding level of crucian carp.

关 键 词：淇河鲫 IL-8 克隆 组织表达 

分 类 号：Q786[生物学—分子生物学] S917.4[农业科学—水产科学]