斜带石斑鱼IFN-γ基因的克隆与表达分析  

Molecular characterization and expression of interferon gamma(IFN-γ) in orange-spotted grouper(Epinephelus coioides)

作　　者：黄贝[1] 陈善楠[1] 黄文树[2] 聂品[1] 

机构地区：[1]中国科学院水生生物研究所,淡水生态与生物技术国家重点实验室,湖北武汉430072 [2]集美大学水产学院,福建厦门361021

出　　处：《中国水产科学》2013年第2期269-275,共7页Journal of Fishery Sciences of China

基　　金：国家自然科学基金委与广东省联合基金资助项目(U0631010);国家“863”计划项目(2012AA092201)

摘　　要：IFN-γ在天然免疫和适应性免疫应答中均发挥着重要的作用。由于低等脊椎动物IFN-γ与哺乳动物IFN-γ相似性非常低,硬骨鱼类的IFN-γ近年来才得到鉴定。本研究报道了斜带石斑鱼(Epinephelus coioides)的IFN-γ基因,并对其表达进行了分析。斜带石斑鱼IFN-γcDNA全长为867 bp,编码200个氨基酸。其基因组全长为5 104 bp,由4个外显子、3个内含子组成。氨基酸序列比对显示斜带石斑鱼IFN-γ与高等脊椎动物IFN-γ的序列相似性较低,仅为32.5%～39%,与其他硬骨鱼类的IFN-γ相似性为38.0%～68.5%。序列分析结果显示,斜带石斑鱼IFN-γ分子C末端中存在IFN-γ的特征序列以及核定位基序。二级结构分析结果显示斜带石斑鱼IFN-γ序列中包含6个α螺旋结构,其排列与高等脊椎动物的IFN-γ类似。应用荧光定量PCR检测了IFN-γ基因在Poly I:C诱导后的表达变化。发现在肾、脾、鳃、头肾、皮肤和胸腺组织中,Poly I:C能显著诱导IFN-γ的表达,在胸腺中上调倍数最高,其次为头肾、脾、肾、皮肤和鳃,而在脑和心脏组织中没有显著变化。据此推测,IFN-γ在斜带石斑鱼抵抗病毒感染的过程中起有十分重要的作用。IFN-y is a type II interferon that plays an important role in both innate and cell-mediated immune re- sponses. Because of the low similarity with their mammalian homologues, IFN-y in teleost fish has only recently been identified. In this study, the cDNA sequence encoding the IFN-y molecule was cloned from the or- ange-spotted grouper, Epinephelus coioides, and its gene organization was revealed. Its expression in response to Poly I：C stimulation was also examined. The IFN-y cDNA sequence consists of 867 bp, which encodes a putative protein of 200 amino acids. The gene is 5-kb long and contains four exons, which are comparable in size with IFN-7 genes in other vertebrates. This IFN-y was only 32.5%-39% similar to its counterparts in higher vertebrates and 38.0%-68.5% similar to those in other teleost fish. Sequence analysis revealed that the IFN-y in the or- ange-spotted grouper contains the typical IFN-y motif and a conserved nuclear localization site in the C-terminal end and displays an alpha-helix structure similar to mammalian IFN-y. Real time PCR analysis results showed that the expression of IFN-7 increased in the thymus, head kidney, spleen, kidney, skin, and gills following the poly I：C stimulation, indicating its role against viruses in orange-spotted grouper.

关 键 词：Ⅱ型干扰素 IFN-Γ 基因表达 斜带石斑鱼 

分 类 号：S941[农业科学—水产养殖]